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Technique development of attractant test for Chrysomya bezziana in laboratory and semi-field conditions Wardhana, April H; ., Sukarsih
Indonesian Journal of Animal and Veterinary Sciences Vol 9, No 1 (2004)
Publisher : Indonesian Animal Sciences Society

Swormlure (SL-2), synthetic attractant for the New World Screwworm Fly (NWSF), Cochliomya hominivorax, have been developed and used in the America. The effectiveness of swormlure in attracting the Old World Screwworm Fly (OWSF), Chrysomya bezziana is not well defined. The aim of the study was to provide suitable condition of the attractant in trapping the higher number at the OWSF in laboratory (cage assay) and semi-field (room assay) conditions. The cage assay to screen responses olfactory stimuli of OWSF was developed to asses the fly responses to lights, exhaust fan (on or off), the fliesâ physiological status and whether there was any bias between cages or trap positions. Modifications were made to provide suitable physical and environmental conditions for candidate attractant. These included darkening all windows with paper, the construction of support for the fly cages and installation of additional lights centred above the fly cages. The room assay was used as an intermediate step between the cage assay and the field experiment. The number of entered flies into the trap indicated flies respond to SL-2. The data of cage assay was analysed by ANOVA and data of room assay was analysed by T test (5%). The results showed that standard experimental conditions for the cage assay: two lights above the cages on and the central lights off, covering fluorescent lights with oil paper, the jar trap positions on the centre line parallel to the lights and exhaust fan was turned off (no air flow) during the session but was turned on in between sessions to reduce the odour from SL-2 in laboratory (p>0.05). The standard experimental conditions for the room assay used four fluorescents tubes, exhaust fan turned off during the replicates but turned on after replicated 3 and 6 for 15 minutes. Yellow half-size sticky was used as standard target (p>0.05). Â Key words: Swormlure, SL-2, attractant, Chrysomya bezziana

Technique development of attractant test for Chrysomya bezziana in laboratory and semi-field conditions April H Wardhana; Sukarsih .
Jurnal Ilmu Ternak dan Veteriner Vol 9, No 1 (2004): MARCH 2004
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Swormlure (SL-2), synthetic attractant for the New World Screwworm Fly (NWSF), Cochliomya hominivorax, have been developed and used in the America. The effectiveness of swormlure in attracting the Old World Screwworm Fly (OWSF), Chrysomya bezziana is not well defined. The aim of the study was to provide suitable condition of the attractant in trapping the higher number at the OWSF in laboratory (cage assay) and semi-field (room assay) conditions. The cage assay to screen responses olfactory stimuli of OWSF was developed to asses the fly responses to lights, exhaust fan (on or off), the flies’ physiological status and whether there was any bias between cages or trap positions. Modifications were made to provide suitable physical and environmental conditions for candidate attractant. These included darkening all windows with paper, the construction of support for the fly cages and installation of additional lights centred above the fly cages. The room assay was used as an intermediate step between the cage assay and the field experiment. The number of entered flies into the trap indicated flies respond to SL-2. The data of cage assay was analysed by ANOVA and data of room assay was analysed by T test (5%). The results showed that standard experimental conditions for the cage assay: two lights above the cages on and the central lights off, covering fluorescent lights with oil paper, the jar trap positions on the centre line parallel to the lights and exhaust fan was turned off (no air flow) during the session but was turned on in between sessions to reduce the odour from SL-2 in laboratory (p>0.05). The standard experimental conditions for the room assay used four fluorescents tubes, exhaust fan turned off during the replicates but turned on after replicated 3 and 6 for 15 minutes. Yellow half-size sticky was used as standard target (p>0.05). Key words: Swormlure, SL-2, attractant, Chrysomya bezziana

In vitro study of larvacidal effect of Chrysomya bezziana between atsiri oil (Piper betle leaf) from Sri Lanka and Bogor April H Wardhana; S Muharsini; S Santosa; L.S.R Arambewela; S.P.W Kumarasinghe
Jurnal Ilmu Ternak dan Veteriner Vol 15, No 4 (2010): DECEMBER 2010
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Treatment of myiasis caused by the larvae of Chrysomya bezziana resulted variation results. The aim of the experiment are to compare larvacidal effect of atsiri oil of Piper betle leaf originated from Sri Lanka and Bogor agains Chrysomya bezziana larvae and to identify atsiri oil using Thin Layer Chromatography (TLC). Silica gel plate GF254 with toluena and ethyl ecetate as diluents was used in TLC. L1 and L2 of Chrysomya bezziana larvae were used for in vitro assay using agar plate contained atsiri oil of three concentrations ie. 2, 3 and 4%. Asuntol 1% and aquadest sterile were used as positive and negative control, respectively. Ten larvae were used in five replicates. The larvae were observed every 30 minutes for four hours. Larval mortality were counted and probit analysed using POLO-PC software, therefore the lethal concentration (LC50 and LC95) and lethal time (LT50 and LT95) were defined. The results showed there was a difference on migration distance between atsiri oil from Sri Lanka and Bogor on nRf 79-88 analysed using TLC. In vitro assay showed that both lethal concentration and lethal time of atsiri oil from Sri Lanka was higher than from Bogor. This results indicated that larvacidal effect of atsiri oil from Bogor was stronger than atsiri oil from Sri Lanka in in vitro both on L1 and L2. Key Words: Piper Bitle Leaf, Atsiri Oil, Chrysomya bezziana, Sri Lanka, Bogor

Characterization of Bacillus thuringiensis isolat from several areas in Java and South Sulawesi for biological control of myasis, Chrysomya bezziana S Muharsini; April H Wardhana; H Rijzaani; B Amirhusein
Jurnal Ilmu Ternak dan Veteriner Vol 8, No 4 (2003): DECEMBER 2003
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Bacillus thuringiensis is a species of bacteria which has potential use for biological control of parasite. The aim of the study is to characterize the local isolate of B. thuringiensis using PCR technique. Primer of cry gen was used in the study. Two hundred and twenty seven soil samples were collected from several areas in Java Island and South Sulawesi Province. Samples were obtained from surrounding soils of animal housing and paddocks. Samples were inoculated in Nutrient Agar and the suspect of bacilli were then cultured in Luria Bertani acetic buffer. Seventy-three pure isolates of Bacillus thuringiensis were obtained. Thirty-five isolates from Bogor, Yogyakarta and Majalengka regencies were amplified using PCR and consisted of a B. thuringiensis sporulation gene which has a 850 bp band in an agarose gel. Amplification using cryIVA,B primers demonstrated four isolates contain a faint band of 800 bp and 12 isolates consist of a distinctive band of 300-400 bp. The results showed that several B. thuringiensis isolates containing a cry IV gen and those isolates are needed to be tested as a biological control agent for Chrysomya bezziana. Key words: Bacillus thuringiensis, Chrysomya bezziana, biological control, cry IV gen

Reliable preservation methods of mitochondrial DNA analysis for larvae and adult flies of Chrysomya bezziana (Diptera: Calliphoridae). April H Wardhana; S Muharsini; Suhardono .
Jurnal Ilmu Ternak dan Veteriner Vol 8, No 4 (2003): DECEMBER 2003
Publisher : Indonesian Center for Animal Research and Development (ICARD)

The Old World Screwworm fly, Chrysomya bezziana is known as a myiasis agent in Indonesia. The aim of the study is to establish a reliable preservation methods for molecular analysis of insect. Cytochrom b (CB) and Cytochrome Oxidase sub unit I (COI) of mitochondrial DNA were used as markers. Drying method was used to preserve adult flies, while the larvae were kept in 10% formalin and 80% ethanol with or without soaked using hot water prior preservation. Two samples of each preservation were tested. Four primers were used to amplify samples using Polymerase Chain Reaction (PCR). The PCR product were then purified and sequenced. Sequence data analysis of all samples suggested no differences between all preservation methods except using 10% formalin. Preservation samples using 10% formalin is not suitable for collecting mitochondrial DNA of insect. The best methods was using 80% ethanol with killed using hot water prior preservation. This method was able to extract a stabile mitochondrial DNA. Key words: Chrysomya bezziana, preservation, formalin, ethanol, mitochondrial DNA

The effectivity of Annona squamosa L seeds extracted by diverse organic solvents: water, methanol and hexane against mortality of tick larvae, Boophilus microplus in vitro April H Wardhana; Amir Husein; J Manurung
Jurnal Ilmu Ternak dan Veteriner Vol 10, No 2 (2005): JUNE 2005
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Boophilus microplus is the most important pest in livestock industries. The use of synthetic chemical acaricides is the main method of tick control, however, chemical acaricides are expensive, and they are harmful to environment and cause strain resistance. The aim of study was to investigate the affectivity of Annona squamosa L seeds extracted by diverse organic solvents such as water, methanol and hexane against mortality of Boophilus microplus larvae in vitro. Five hundred and fifty larvae were used in this study and divided into three groups e.g. water (3, 4 and 5% concentration), methanol and hexane extract groups (0.25, 0.50, and 0.75% concentration). Coumaphos (0.50%) was used as a positive control. The larvae were dipped into extract solution for 10 seconds and dried using filter paper. Their mortality was observed from one to five hours. The mortality data were transformed to Abbot formula and analyzed using probit analysis with 95% significant level. This study showed that the active compound of Annona squamosa L seeds had effectively contact toxic property for B. microplus larvae at 5, 0.50, and 0.75% for water, methanol and hexane extractions, respectively. The lethal concentrations of methanol extract (LC50, LC90, and LC95) were lower than hexane extract e.g. 0.32, 0.86, and 1.13%, respectively and for hexane extract were 0.35, 1.11, and 1.54%, respectively at fifth hour. The lethal times of methanol extract on 0.50% concentration were shorter than hexane extract e.g. 3.12 hours (LT50), 5.86 hours (LT90), and 7.00 hours (LT95) and for hexane extract on 0.75% concentration were 3.26 hours (LT50), 6.21 hours (LT90), and 7.45 hours (LT95). Water extract of 5% concentration was effective for traditional farmer in rural area due to easy and cheap method. The lethal concentrations of water extract on fifth hour were 2.02% (LC50), 4.00% (LC90), and 4.85% (LC95) and the lethal time on 5% concentration were 2.54 hours (LT50), 4.13 hours (LT90), and 4.75 hours (LT95). Key Words: Annona squamosa, Boophilus microplus, Water Extract, Methanol, Hexane

icacy test of hexane extract of Annona squamosa L seeds for Chrysomya bezziana larvae growth in vitro April H Wardhana; E Widyastuti; A.W.A Wiratmana; S Muharsini; Darmono .
Jurnal Ilmu Ternak dan Veteriner Vol 9, No 4 (2004): DECEMBER 2004
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Chrysomya bezziana is primer agent causing myiasis in livestock and human throughout Africa to Asia. At present, treatment of myiasis using chemical synthetic insecticide causing environment and livestock production contaminations. The aim of this study was to evaluate the effect of hexane extract of Annona squamosa L seed to kill C. bezziana as botanical insecticide. Three level and method preparation of larvae (L1, L2 and L3) were carried out. Meat blood mixture (MBM) and larval rearing media (LRM) were mixed with hexane extract of A. squamosa L seed and tested to 625 and 750 larvae (L1and L2 respectively), while for L3 625 larvae were soaked in the solution containing the hexane extract. This last preparation was carried out to observe the effect of toxic contact. Each larvae treatment was divided into 5 levels such as negative control using distillation water (PO), given 0.25% hexane extract (P I); 0.50% (P II); 0.75% (P III) and positive control was given with 0.10% Asuntol®. This assay was to know digest toxic effect of hexane extract of A. squamosa L seed for those larvae. Number of 625 instar III larvae (L3) was soaked in the solution containing the hexane extract of A. squamosa L seed for 10 second, placed into vermicullite and incubated on 36oC. This assay was to know contact toxic effect of them. All of larvae were allowed to become pupae. Parameters oberserve were number of larvae death, pupae weight and number of pupae become into fly. The data was analyzed using Anova (5%) and Z test (5%) then smallest significant difference test (BNT 5%). The results showed that 0.50% of hexane extract of A. squamosa L seed was able to decrease pupae weight for L1 and L2 and to cause fail pupae become fly (P<0.05). Instar III larvae (L3) soaked in PI until P III and not effect to pupae weight, pupae become fly and survival of adult flies (P>0.05). Key words: Chrysomya bezziana, myasis, srikaya, A. squamosa L, in vitro

The effect of “patikan kebo”(Euphorbia hirta L) preparations on erythrocyte number, haemoglobin level and haematocrit value of chicken infected with Eimeria tenella April H Wardhana; E Kencanawati; Nurmawati .; Rahmaweni .; C.B Jatmiko
Jurnal Ilmu Ternak dan Veteriner Vol 6, No 2 (2001): JUNE 2001
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Patikan kebo” (Euphorbia hirta L) is a medicinal plant mostly used to treat gastro-intestinal track problems empirically. The aim of this study was to determine the effect of distillation, infusion and extract of “patikan kebo” (Euphorbia hirta L) preparations on erythrocyte number, haemoglobin level and haematocrit value in chickens that were infected by Eimeria tenella. Twenty four Hubbard strain broiler chicken raised from one-day-old until three-week-old were used for this study and divided randomly into 4 groups: P 0 (control), P I (distillation treatment), P II (infusion treatment), and P III (extract treatment). A total of 5.000 oocyst of E. tenella were infected into each chicken and the dose of Euphorbia hirta L was 500 mg/kg Wb. A completely randomized design was used and the data were tested by ANOVA. The result indicated that distillation, infusion and extract preparations of Euphorbia hirta L had the potency to increase erythrocyte number, haemoglobin level and haematocrit value. Key word : Eimeria tenella, Euphorbia hirta L, erythrocyte, haemaoglobin, haematocrit

Identification of volatile compounds from myiasis wounds and its responsesfor Chrysomya bezziana April H Wardhana; Sukarsih .; R Urech
Jurnal Ilmu Ternak dan Veteriner Vol 10, No 1 (2005): MARCH 2005
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Development of attractant for screwworm fly was required in myiasis control on livestock. The purpose of this study is to identify of volatile compounds from myiasis wound infested with Chrysomya bezziana larvae including to assess their responses in both cage and room assays. Both Friesian-Holstein heifer (FH) (animal 1) and Bali cattle (animal 2) were used as myiasis model. The artificial wounds (8-10 cm) were conducted on the rump of both animals and infested with about 200 eggs of C.bezziana. Odours from the infested wound were collected on day 1 and 3 for animal1 and day 3 and 5 for animal 2, post C. bezziana larvae infestation. Two different collection devices were used: firstly, absorption onto Tenax kept in steel tubes, whichwas attached to a collected bowl. The volatile organic compounds were collected from the wound and the surrounding animal hide by flowins the air through the inlet and outlet. Secondly, a solid phase micro extraction (SPME) device was inserted into bowl with passive (no air flow) odour collection. Gass chromatography/mass spectrometry was used to identify volatile compounds from wound. The compounds of the wound on animal 1, collected on day 1, produced only minor quantities of compounds (nonanal, decanal, hexanal and heptanal). Minor components such as DMDS and DMTS were only detected on day 3. The compounds of the wound on animal 2 was more varied and had a peculiar strike-like smell on day 3 and 5. They included indole, phenol, acetone, various sulfides (DMS, DMDS, DMTS), alcohols (butanol, 3-methylbutanol), aldehydes and acids. These compounds were selected and formulated into attractant (B92) then tested in both cage and room assays using SL-2 as control. Respond of flies was analyzed by ANOVA 5% (cage assay) and T test 5% (room assay). The result showed that the fly response to B92 was very low compared to SL-2 in cage assays (P<0.05). The addition of B92 to SL-2 could not increase the catch of flies in the cage assays (SL-2+B92=10:1; 10:3), there was no difference between SL-2 and B92/SL-2 in room assay, the fly response still low (P>0.05). Key Words: Volatile Compound, Myiasis, Wound, Chrysomya bezziana

diversity of myiasis fly, Chrysomya bezziana population in Indonesia based on mitochondrial DNA April H Wardhana; S Muharsini; Widya Asmara
Jurnal Ilmu Ternak dan Veteriner Vol 9, No 2 (2004): JUNE 2004
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Chrysomya bezziana is a fly causing myiasis in most livestock in Indonesia. To date, the genetic diversity of C. bezziana has been argued among researchers. The aim of this study was to analyse genetic diversity of C. bezziana population in Indonesia using mitochondrial cytochrome b gene (cyt b) as a marker. The preliminary study showed that the larvae instar III stadium (L3) was the most appropriate sample for molecular analysis and identification. Twenty-four L3 were collected from cattles, buffalos and horses in Bogor, Makassar and East Sumba. DNA samples were isolated from muscle tissue of the larvae. The fragment of cyt b gene (279 bp) was amplified using primer CB3FC-NINFA. The PCR product was subsequently purified, sequenced and analysed using PAUP version 4. The results showed that C. bezziana from Indonesia was different with those from Asia, basicaly for DNA sequences, however it had similar to those from Papua New Guinea. Two haplotypes were identified i.e. haplotype 6 (Makassar haplotype that similar to those from Papua New Guinea) and haplotype 7 (Bogor haplotype that similar to those from Makassar and East Sumba). Key words: Chrysomya bezziana, genetic diversity, mitochondrial DNA

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