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Papers by Peshala Gunasekara

Research paper thumbnail of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Allergy, Asthma & Clinical Immunology, 2019

BackgroundAllergy toVespa affinisvenom is common in the Asia Pacific region. Venom preparations f... more BackgroundAllergy toVespa affinisvenom is common in the Asia Pacific region. Venom preparations for diagnosis are not commercially available for this species.MethodsThe prominent allergens inV. affinisvenom were identifiedusing immunochemical methods. Use of ImmunoCAP ofVespula vulgariscrude venom/its components and a passive basophil activation test (BAT) in the diagnosis of patients who had anaphylaxis toV. affinisvenom (n = 30) were also accessed. The IgE double-positivity rates (positive to both hornet and honeybee) in ImmunoCAP and the passive BAT were determined.ResultsHigh IgE reactivity was seen with the five allergens inV. affinisvenom; 96% (29/30) for 34 and 24 kDa, 93% (28/30) for 45 kDa and 90% (27/30) reactivity for the 100 and 80 kDa respectively. IgE cross-reactivity was low with ImmunoCAP usingV. vulgarisvenom (43%; 13/30) and Ves v1 (3%; 1/30), but relatively high with Ves v5 (73%; 22/30). All patients (100%) were positive toV. affinisvenom in passive BAT. In Immuno...

Research paper thumbnail of Identification of allergens in coconut milk and oil with patients sensitized to coconut milk in Sri Lanka

Clinical and Molecular Allergy

Background Despite the low prevalence of IgE sensitivity to fresh or boiled coconut milk and coco... more Background Despite the low prevalence of IgE sensitivity to fresh or boiled coconut milk and coconut oil, those may contain allergens of which the clinical significance remains undetermined. This study aimed to identify and compare allergens in fresh coconut milk (FCM), boiled coconut milk (BCM), unrefined wet-processed coconut oil (WPCO), and dry-processed coconut oil (DPCO) using sera from patients with allergy to coconut milk. Methods The study included 18 patients with immediate hypersensitivity to coconut milk, including five who developed anaphylaxis. Sensitization was assessed by skin prick test and ImmunoCAPs using commercially available coconut extracts. Immunoblotting was performed to identify and compare allergen profiles. Results Total sIgE levels and overall IgE reactivity of patients with anaphylaxis were higher compared to patients with allergy. Twelve allergens ranging from 5 to 128 kDa including six novel allergens with 5, 12, 47, 87, 110, and 128 kDa were visualize...

Research paper thumbnail of MOESM1 of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Additional file 1: Figure S1. Comparison of IgE reactivity; Specific IgE to crude venom of V. vul... more Additional file 1: Figure S1. Comparison of IgE reactivity; Specific IgE to crude venom of V. vulgaris and Ves v 5 (P = 0.02 in Mann–Whitney U test) Horizontal bars indicate the mean of specific IgE quantity to either V. vulgaris venom or Ves v 5 and the dotted line represent the Phadia ImmunoCAP cut off level 0.1 kUA/l. Figure S2. Generation of passive immune donor basophils; (a) membrane bound IgE on donor basophils (b) removal of donor IgE by lactic acid treatment and (c) reattachment of patient IgE; top—scatter diagram and bottom—intensity diagram.

Research paper thumbnail of Comparison of two rapid test kits with real time polymerase chain reaction for early diagnosis of dengue in Sri Lanka

Journal of Immunoassay and Immunochemistry, 2021

ABSTRACT Dengue is among the deadliest insect-borne diseases circulating in Sri Lanka. Most of th... more ABSTRACT Dengue is among the deadliest insect-borne diseases circulating in Sri Lanka. Most of the infections that are diagnosed early are manageable. However, delays in diagnosis may cause fatalities. We evaluated the dengue NS1 antigen card and NS1 SD kit for early diagnosis of dengue using samples from 116 RT-PCR-positive patients admitted within 5 days of the fever onset. RT-PCR tests were performed as standard tests. IgM and IgG ELISA tests were carried out to identify primary and secondary infections. Of the 116 patients who tested positive for dengue using PCR, 48 were positive using NS1 antigen card and 45 were positive using NS1 SD. Patients with 100 copies or higher viral load showed a higher sensitivity in both antigen card and NS1 SD. Of 34 primary infections evaluated, 23 were positive by NS1 antigen card, while the positivity was 21 by NS1 SD. Of the 30 secondary infections evaluated, 15 were positive by NS1 antigen card while 14 by NS1 SD. Our findings showed that while the rapid tests are convenient and much easier to use than PCR, they are less sensitive and need improvement. Until then, clinical diagnosis should have more emphasis on the early diagnosis of dengue.

Research paper thumbnail of 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

Toxicon, 2017

Diagnostic and therapeutic reagents are unavailable for anaphylaxis arising from stings by Apis d... more 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.

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

Clinical and Molecular Allergy

Background Allergy to Apis dorsata (Giant Asian Honeybee) venom is the commonest insect allergy i... more 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 t...

Research paper thumbnail of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Allergy, Asthma & Clinical Immunology, Dec 1, 2019

BackgroundAllergy to Vespa affinis venom is common in the Asia Pacific region. Venom preparations... more BackgroundAllergy to Vespa affinis venom is common in the Asia Pacific region. Venom preparations for diagnosis are not commercially available for this species.MethodsThe prominent allergens in V. affinis venom were identifiedusing immunochemical methods. Use of ImmunoCAP of Vespula vulgaris crude venom/its components and a passive basophil activation test (BAT) in the diagnosis of patients who had anaphylaxis to V. affinis venom (n = 30) were also accessed. The IgE double-positivity rates (positive to both hornet and honeybee) in ImmunoCAP and the passive BAT were determined.ResultsHigh IgE reactivity was seen with the five allergens in V. affinis venom; 96% (29/30) for 34 and 24 kDa, 93% (28/30) for 45 kDa and 90% (27/30) reactivity for the 100 and 80 kDa respectively. IgE cross-reactivity was low with ImmunoCAP using V. vulgaris venom (43%; 13/30) and Ves v1 (3%; 1/30), but relatively high with Ves v5 (73%; 22/30). All patients (100%) were positive to V. affinis venom in passive BAT. In ImmunoCAP, a high double-positivity rate (76%; 23/30) was detected while no double-positivity was detected in passive BAT.ConclusionsHigh IgE reactivity for five allergens of V. affinis points to the potential of using these allergens in component resolved diagnosis (CRD). The passive BAT has shown its importance as a promising diagnostic tool with high accuracy. It would be particularly useful in cases with doubtful double-positive results of other diagnostic tests.

Research paper thumbnail of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Allergy, Asthma & Clinical Immunology, 2019

BackgroundAllergy toVespa affinisvenom is common in the Asia Pacific region. Venom preparations f... more BackgroundAllergy toVespa affinisvenom is common in the Asia Pacific region. Venom preparations for diagnosis are not commercially available for this species.MethodsThe prominent allergens inV. affinisvenom were identifiedusing immunochemical methods. Use of ImmunoCAP ofVespula vulgariscrude venom/its components and a passive basophil activation test (BAT) in the diagnosis of patients who had anaphylaxis toV. affinisvenom (n = 30) were also accessed. The IgE double-positivity rates (positive to both hornet and honeybee) in ImmunoCAP and the passive BAT were determined.ResultsHigh IgE reactivity was seen with the five allergens inV. affinisvenom; 96% (29/30) for 34 and 24 kDa, 93% (28/30) for 45 kDa and 90% (27/30) reactivity for the 100 and 80 kDa respectively. IgE cross-reactivity was low with ImmunoCAP usingV. vulgarisvenom (43%; 13/30) and Ves v1 (3%; 1/30), but relatively high with Ves v5 (73%; 22/30). All patients (100%) were positive toV. affinisvenom in passive BAT. In Immuno...

Research paper thumbnail of Identification of allergens in coconut milk and oil with patients sensitized to coconut milk in Sri Lanka

Clinical and Molecular Allergy

Background Despite the low prevalence of IgE sensitivity to fresh or boiled coconut milk and coco... more Background Despite the low prevalence of IgE sensitivity to fresh or boiled coconut milk and coconut oil, those may contain allergens of which the clinical significance remains undetermined. This study aimed to identify and compare allergens in fresh coconut milk (FCM), boiled coconut milk (BCM), unrefined wet-processed coconut oil (WPCO), and dry-processed coconut oil (DPCO) using sera from patients with allergy to coconut milk. Methods The study included 18 patients with immediate hypersensitivity to coconut milk, including five who developed anaphylaxis. Sensitization was assessed by skin prick test and ImmunoCAPs using commercially available coconut extracts. Immunoblotting was performed to identify and compare allergen profiles. Results Total sIgE levels and overall IgE reactivity of patients with anaphylaxis were higher compared to patients with allergy. Twelve allergens ranging from 5 to 128 kDa including six novel allergens with 5, 12, 47, 87, 110, and 128 kDa were visualize...

Research paper thumbnail of MOESM1 of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Additional file 1: Figure S1. Comparison of IgE reactivity; Specific IgE to crude venom of V. vul... more Additional file 1: Figure S1. Comparison of IgE reactivity; Specific IgE to crude venom of V. vulgaris and Ves v 5 (P = 0.02 in Mann–Whitney U test) Horizontal bars indicate the mean of specific IgE quantity to either V. vulgaris venom or Ves v 5 and the dotted line represent the Phadia ImmunoCAP cut off level 0.1 kUA/l. Figure S2. Generation of passive immune donor basophils; (a) membrane bound IgE on donor basophils (b) removal of donor IgE by lactic acid treatment and (c) reattachment of patient IgE; top—scatter diagram and bottom—intensity diagram.

Research paper thumbnail of Comparison of two rapid test kits with real time polymerase chain reaction for early diagnosis of dengue in Sri Lanka

Journal of Immunoassay and Immunochemistry, 2021

ABSTRACT Dengue is among the deadliest insect-borne diseases circulating in Sri Lanka. Most of th... more ABSTRACT Dengue is among the deadliest insect-borne diseases circulating in Sri Lanka. Most of the infections that are diagnosed early are manageable. However, delays in diagnosis may cause fatalities. We evaluated the dengue NS1 antigen card and NS1 SD kit for early diagnosis of dengue using samples from 116 RT-PCR-positive patients admitted within 5 days of the fever onset. RT-PCR tests were performed as standard tests. IgM and IgG ELISA tests were carried out to identify primary and secondary infections. Of the 116 patients who tested positive for dengue using PCR, 48 were positive using NS1 antigen card and 45 were positive using NS1 SD. Patients with 100 copies or higher viral load showed a higher sensitivity in both antigen card and NS1 SD. Of 34 primary infections evaluated, 23 were positive by NS1 antigen card, while the positivity was 21 by NS1 SD. Of the 30 secondary infections evaluated, 15 were positive by NS1 antigen card while 14 by NS1 SD. Our findings showed that while the rapid tests are convenient and much easier to use than PCR, they are less sensitive and need improvement. Until then, clinical diagnosis should have more emphasis on the early diagnosis of dengue.

Research paper thumbnail of 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

Toxicon, 2017

Diagnostic and therapeutic reagents are unavailable for anaphylaxis arising from stings by Apis d... more 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 (&amp;amp;gt;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.

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

Clinical and Molecular Allergy

Background Allergy to Apis dorsata (Giant Asian Honeybee) venom is the commonest insect allergy i... more 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 t...

Research paper thumbnail of Diagnosis of Vespa affinis venom allergy: use of immunochemical methods and a passive basophil activation test

Allergy, Asthma & Clinical Immunology, Dec 1, 2019

BackgroundAllergy to Vespa affinis venom is common in the Asia Pacific region. Venom preparations... more BackgroundAllergy to Vespa affinis venom is common in the Asia Pacific region. Venom preparations for diagnosis are not commercially available for this species.MethodsThe prominent allergens in V. affinis venom were identifiedusing immunochemical methods. Use of ImmunoCAP of Vespula vulgaris crude venom/its components and a passive basophil activation test (BAT) in the diagnosis of patients who had anaphylaxis to V. affinis venom (n = 30) were also accessed. The IgE double-positivity rates (positive to both hornet and honeybee) in ImmunoCAP and the passive BAT were determined.ResultsHigh IgE reactivity was seen with the five allergens in V. affinis venom; 96% (29/30) for 34 and 24 kDa, 93% (28/30) for 45 kDa and 90% (27/30) reactivity for the 100 and 80 kDa respectively. IgE cross-reactivity was low with ImmunoCAP using V. vulgaris venom (43%; 13/30) and Ves v1 (3%; 1/30), but relatively high with Ves v5 (73%; 22/30). All patients (100%) were positive to V. affinis venom in passive BAT. In ImmunoCAP, a high double-positivity rate (76%; 23/30) was detected while no double-positivity was detected in passive BAT.ConclusionsHigh IgE reactivity for five allergens of V. affinis points to the potential of using these allergens in component resolved diagnosis (CRD). The passive BAT has shown its importance as a promising diagnostic tool with high accuracy. It would be particularly useful in cases with doubtful double-positive results of other diagnostic tests.