Validation of a subjective counting method for a horn flies (Haematobia irritans irritans) (Diptera: Muscidae) population in a cattle herd

A trained observer direct count method to measure horn fly population was evaluated to determine the reliability (inter-observer agreement) and its validity when compared with a "gold standard" method (video film). All the counts were performed with the animals restrained in a chute in a single herd. A direct count of horn flies by each of two observers on opposite sides of the animal was made. In addition a videotape recording of the counting surface on each animal was made. Horn flies were counted on 80 cows in the morning twice monthly from 22 October 1999 to 24 March 2000. The correlation between observer counts was high as was the correlation between observer counts and counts made from a videotape recording. Direct counts by trained observers were highly reliable and had good validity. However, videotape recording can be useful when the horn fly population is high, because it is faster in the field than the direct count method. The results of this study suggest that the direct count method can be used with confidence under field conditions.     

Population dynamics of horn fly, Haematobia irritans irritans (L.) (Diptera: Muscidae), on Hereford cattle in Uruguay

Abundance of adult horn flies, Haematobia irritans irritans (L.), was monitored on 25 untreated Hereford cows in Tacuarembó Department, Uruguay, during three consecutive grazing seasons, from October 1999 to May 2002. The population showed a variable pattern of abundance during three years, with peaks in late summer-early fall of each year. Adult flies were continuously present, although in very low numbers in intervening winters. Numbers of flies per cow rarely exceeded a reference level of 200 flies per animal during the grazing season. Degree-day calculations indicated that approximately 12 generations were possible each year. Time series analysis of mean densities among consecutive generations indicated that population growth was governed by simple, direct density-dependence, with additional effects of seasonally varying weather. Response surface regressions confirmed that intergenerational growth was inversely related to mean density, and directly related to temperature. Stochastic simulations with the response surface model suggested that within the range of temperatures observed in our study, horn fly populations on Hereford cattle will tend toward densities of approximately 150 flies per animal in summer, and exceed a nominal level of 200 flies per cow one or more times in about 65 of every 100 grazing seasons.     

Acetylcholinesterase mutation in diazinon-resistant Haematobia irritans (L.) (Diptera: Muscidae)

Acetylcholinesterase (AChE) cDNA from individual field-collected diazinon-resistant horn flies was amplified by RT-PCR. Sequencing of the amplification products revealed that 8/12 of the diazinon-resistant horn flies contained a point mutation previously associated with resistance to organophosphates in house flies and Drosophila, strongly suggesting that this cDNA encodes the AChE that is the target site for organophosphate (OP) pesticide. The point mutation (G262A) resulted in a shift from glycine to alanine in the mature HiAChE amino acid sequence at position 262. Allele-specific PCR and RLFP assays were developed to diagnose the presence or absence of the G262A mutation in individual flies. Use of the allele-specific assays each demonstrated the presence of the G262A mutation in 10 of 12 individual field-collected flies, demonstrating higher sensitivity than direct sequencing of RT-PCR amplification products. The G262A mutation was found in additional fly populations previously characterized as OP-resistant, further supporting that this AChE is the target site for OP pesticide. The allele-specific assay is a useful tool for quantitative assay of the resistance allele in horn fly populations.     

Chlorfenapyr ear tags to control Haematobia irritans (L.) (Diptera: Muscidae) on cattle

The efficacy of ear tags containing 30% chlorfenapyr (total tag weight=13g) to control natural Haematobia irritans (L.) infestations was evaluated for Holstein heifers in Rafaela, province of Santa Fe, Argentina. A group of heifers (TG) was treated with two ear tags (one tag per ear). A control group (ACG) was maintained in a paddock adjacent to the TG paddock and, a distant control group (DCG) was maintained 700m apart from the other groups. From day 4 to day 98 after treatment, H. irritans infestations of ACG were significantly higher (P<0.05, test of Kruskal-Wallis) than the corresponding infestation of TG, but significantly lower than infestation in DCG, probably due to the proximity of TG. The chlorfenapyr control period, with an efficacy higher than 90% to reduce horn fly populations, lasted for 9 weeks when TG infestation was compared to fly numbers in ACG, but increased to 12 weeks in comparison to DCG. The results of this study show that ear tags impregnated with chlorfenapyr are a useful alternative to pyrethroids and organo-phosphate compounds for horn fly control.     

骆驼斯氏副柔线虫病传播媒介西方角蝇和截脉角蝇各虫态的系统形态学研究

本研究旨在完善西方角蝇和截脉角蝇各虫态的系统形态学资料,为野外采集两种角蝇各虫态样本的分类鉴定提供理论依据,进而为研究斯氏副柔线虫在角蝇体内发育过程,以及制定防控骆驼斯氏副柔线虫病的有效措施等奠定基础。通过实验室人工饲养角蝇,收集西方角蝇和截脉角蝇的卵、各龄幼虫、蛹和成蝇,详细观察并描述两种角蝇各虫态的形态结构,以明确两种角蝇各虫态之间的形态学差异。除西方角蝇和截脉角蝇1龄幼虫外,两种角蝇的卵、2龄幼虫、3龄幼虫、蛹和成蝇都存在可识别的形态学差异。本研究详细描述了西方角蝇和截脉角蝇各虫态的形态特征,并确定了多处可用于区分两种角蝇卵、2龄幼虫、3龄幼虫、蛹和成蝇的形态特征差异。     

Toxicity of cypermethrin and diazinon to Haematobia irritans (Diptera: Muscidae) in its American southern range

Synthetic pyrethroids are considered currently less effective than organophosphate insecticides for controlling Haematobia irritans on cattle in its American southern range. To test the validity of this hypothesis, horn fly populations from 95 cattle herds in central-northern Argentina and southern Brazil were evaluated to determine 50% lethal concentrations (LC(50)) to cypermethrin and diazinon, respectively. Horn flies were exposed to filter papers and glass vials treated with cypermethrin and diazinon, respectively, and LC(50) values were determined using probit analysis. Two and 78 fly populations showed LC(50) values indicating susceptibility and resistance to cypermethrin, respectively, in comparison to historical LC(50) values obtained when the horn fly populations were susceptible to cypermethrin. Five fly populations had an undetermined resistance status to cypermethrin. The diazinon tests showed no obvious indications of H. irritans resistance. Thirty nine percent of farmers still rely on pyrethroids to control horn fly populations on cattle which represents a decreasing proportion from a few years ago when horn fly control was carried out using pyrethroids almost exclusively. Organophosphates could provide control for horn flies resistant to pyrethroids; however, the reliance on chemicals alone to control H. irritans is not considered a proper approach for the future.     

A study of relative horn fly,Haematobia irritans (Diptera: Muscidae), abundance on Holstein steers and steers of two Holstein crosses

A study was performed to determine if the number of horn fly (Haematobia irritans) adults differ significantly on Holstein (black and white coat color), Holstein x Holstein Friesian (black and white coat color) and Holstein x Jersey (black coat color) steers, 10-12 months old at the onset of the study. All steers were run together on lucerne paddocks and the number of flies counted at 30-day intervals from September 2000 to August 2001. No significant differences (P > 0.05, test of Kruskal-Wallis) were found in fly numbers, even in the period April-May 2001 when the infestation reached its peak. We were unable to demonstrate that coat color influenced horn fly abundance in the present study. It appears that none of the biotypes evaluated had any advantage for natural control of H. irritans.     

Cypermethrin pour-on synergized with piperonyl butoxide: effects on Haematobia irritans (Diptera: Muscidae) natural populations resistant to cypermethrin.

Development of pyrethroid resistance in Haematobia irritans in Santa Fe province, Argentina, resulted in an increased use of pyrethroid insecticides, probably due to lack of suitable alternative treatments. We explored the efficacy of mixtures of cypermethrin and piperonyl butoxide (PBO) against pyrethroid-resistant H. irritans. Groups of 25 Holstein cows each, naturally infested with cypermethrin resistant H. irritans were assigned to treated or control groups in April, September, October and December 1997. Cattle in treated groups were medicated with pour-on oil formulations of 5% cypermethrin (dose = 4 mg per kg of body weight) with 5% or 10% PBO in April, and with a mixture containing 5% of both components thereafter. Efficacy was tested for 21 days after treatment. A treatment of 5% cypermethrin pour-on without PBO was evaluated in October 1997. Samples of horn flies were obtained before September, October and December treatments and exposed for 2 h to filter papers impregnated with different cypermethrin concentrations to determine the 50% lethal concentration (LC50). No difference in efficacy was found between cypermethrin pour-on formulations with 5% or 10% of PBO (more than 94% efficacy on day 21 after treatment). Efficacy of 5 % cypermethrin-5% PBO mixture decreased rapidly in the successive treatments (less than 40% efficacy was observed on day 21 after the December treatment), and the period after treatment with an efficacy higher than 95% was 14 days for the treatment carried out in April, 10 days in September; 7 days for the treatment performed in October and 4 days for the December treatment. The LC50 of cypermethrin was 36.6 microg per cm2 in September and increased to 116.6 and 226.1 microg per cm2 in October and December, respectively. It is concluded that the addition of PBO to cypermethrin did not provide a treatment that would give a long term control of pyrethroid resistant-horn flies.     

Genome‐wide association study for feedlot average daily gain in Nellore cattle (Bos indicus)

The genome‐wide association study (GWAS) results are presented for average daily gain (ADG) in Nellore cattle. Phenotype of 720 male Bos indicus animals with information of ADG in feedlots and 354 147 single‐nucleotide polymorphisms (SNPs) obtained from a database added by information from Illumina Bovine HD (777 962 SNPs) and Illumina BovineSNP50 (54 609) by imputation were used. After quality control and imputation, 290 620 SNPs remained in the association analysis, using R package Genome‐wide Rapid Association using Mixed Model and Regression method GRAMMAR‐Gamma. A genomic region with six significant SNPs, at Bonferroni‐corrected significance, was found on chromosome 3. The most significant SNP (rs42518459, BTA3: 85849977, p = 9.49 × 10^?8) explained 5.62% of the phenotypic variance and had the allele substitution effect of ?0.269 kg/day. Important genes such as PDE4B, LEPR, CYP2J2 and FGGY are located near this region, which is overlapped by 12 quantitative trait locus (QTLs) described for several production traits. Other regions with markers with suggestive effects were identified in BTA6 and BTA10. This study showed regions with major effects on ADG in Bos indicus in feedlots. This information may be useful to increase the efficiency of selecting this trait and to understand the physiological processes involved in its regulation.     

Blood Supply in the Tongue of Nellore Bos indicus (Linnaeus, 1758)

Aspects of the vascularization of the tongue of Nellore Bos indicus ( Linnaeus, 1758) were evaluated through the vascular injection technique (with latex-type Neoprene 450, Du Pont do Brasil S.A. and Sulvinil coloring, Glassurit), fixed in formaldehyde at 7% and dissected with magnifying glass. The material was collected at Goiás Carnes Freezer Warehouse in Goiania, Goias. It was found that the deep lingual artery penetrated the lower lateral region of the prominence of the dorsal area of the tongue, advancing rostrally between the hyoid bone and the hypoglossal muscle. In the intravisceral initial third, the artery represents the deep, sinuous continuation branch of the lingual artery, in which path the sublingual artery was stressed in the ventral plan. Then, the artery deepened in the interior of the hypoglossal muscles and genioglossal, supplying dorsal branches (from three to nine) for the lingual torus; from one to five dorsal collateral branches for the lingual cavity; and one or two ventral collateral branches. At the lingual apex, the artery undergoes bifurcation supplying collateral, dorsal and ventral branches with anastomoses between the parallel vessels in the same antimere and between branches of lower caliber hierarchy between opposite antemeres. The large presence of microvessels indicates a significant blood supply to the organ. These results, in comparison with those found in literature, suggest a peculiar vascular pattern for this cattle breed of Indian origin.     

Association of the kdr and superkdr sodium channel mutations with resistance to pyrethroids in Louisiana populations of the horn fly, Haematobia irritans irritans (L.)

Pyrethroid resistance in three horn fly populations in Louisiana was monitored by weekly fly counts, filter paper bioassays, and diagnostic PCR assays for the presence of pyrethroid resistance-associated mutations in the sodium channel gene coding region. The PCR assay for the knockdown resistance (kdr) and superkdr sodium channel mutations was used to determine the frequency of the target site insensitivity mechanism in the populations of horn flies, which possessed varying degrees of insecticide resistance. The bioassays and frequency of homozygous-resistant (RR) kdr genotypes were relative predictors of the fly control subsequently observed. Flies exposed to filter paper impregnated with a discriminating concentration of one of four different insecticides were collected when 50% mortality was estimated. Genotypes for the dead flies and the survivors were determined by the PCR assay. The results of the PCR assays indicated that the genotype at the kdr locus of the flies exposed to the two pyrethroids had an effect upon whether the flies were considered to be alive or dead at the time of collection. The kdr genotype of flies exposed to either diazinon or doramectin was unrelated to whether the flies were considered to be alive or dead, except for a single comparison of flies exposed to diazinon. When possible interactions of the kdr and superkdr mutations were compared, we found that there were no associations with the response to diazinon and doramectin. For one location, there were no survivors of the 75 flies with the SS-SS (superkdr-kdr) homozygous susceptible wild type genotype exposed to pyrethroids, while there were survivors in all of the other five genotypes. The SS-RR genotype flies were more susceptible to the pyrethroids than the SR-RR flies, but that was not the case for exposure to diazinon or doramectin. In the St. Joseph population, there were an adequate number of flies to demonstrate that the SS-SR genotype was more susceptible to pyrethroids than the SS-RR and that flies with the SR-SR genotype were more susceptible to pyrethroids than the flies with the SR-RR genotype.     

Spermatogenesis in Bali cattle (Bos sondaicus) and hybrids with Bos indicus and Bos taurus

Semen quality, testis size and efficiency of sperm production in Bali cattle (Bos sondaicus) and hybrids with Bos indicus and Bos taurus were determined. Mean (+/- SEM) daily sperm production per gram of testis parenchyma (DSPG) in six purebred Bali bulls was 12.2 +/- 0.7 x 10(6). F1 B sondaicus cross B taurus bulls and F1 B sondaicus cross B indicus bulls were sterile. Spermatogenesis was arrested at the late primary spermatocyte stage. In 11 B sondaicus cross B indicus crosses, mean DSPG was lower than in the purebred B sondaicus, although four (one 1/4 B sondaicus, one 3/4 B sondaicus, one 5/8 B sondaicus inter se and one 3/8 B sondaicus inter se) exhibited DSPG levels similar to the foundation stock. Semen from those crossbreeds which exhibited complete spermatogenesis was more variable in terms of spermatozoal concentration, percentage of spermatozoa exhibiting progressive motility and levels of spermatozoal abnormalities. In crossbreeds where sperm production was reduced or absent, there was seminiferous epithelial dysfunction, manifested as an increased frequency of degenerative late pachytene and diplotene primary spermatocytes and germinal cells occurring later in the cycle, or in extreme cases, as complete arrest of spermatogenesis at the late primary spermatocyte stage.     

Pharmacokinetics of diminazene in female Boran (Bos indicus) cattle

The disposition kinetics and bioavailability of diminazene in five healthy heifers were determined after single intravenous (i.v.) and intramuscular (i.m.) administration of the drug in sequence with a wash-out period between administrations of 6 weeks. Intact diminazene in plasma, whole blood and urine samples was analysed using high-performance liquid chromatography. Nonlinear regression analysis of the i.v. and i.m. data indicated that, for either route, the plasma disappearance curves of diminazene were best described by triexponential equations. The i.v. bolus was followed by rapid and biphasic distribution with half-life values of 0.04 h and 0.58 h, V_d(_ss) was 1.91 ± 0.42 1/kg, elimination half-life was 31.71 h while CI averaged 1.74 ± 0.40 ml/min/kg. Within 30 min of the i.v. dose, the erythrocyte/plasma partition ratio of diminazene was 0.30 ± 0.15. Diminazene was rapidly absorbed following i.m. administration; t _½k_a was 0.60 h. C_max, 4.68 ± 1.12 μg/ml, was attained in 10–15 min and systemic availability was 102.42 ± 7.25%. The half-life of the terminal disappearance phase was 145.48 h. About 8.26% of the i.m. dose was excreted intact in the urine within the first 24 h of treatment. In vitro , diminazene was bound to bovine plasma albumin to the extent of 38.01–91.10%.     

Thrombostasin isoform frequency in a Central Texas field population of the horn fly, Haematobia irritans

Thrombostasin is an anti-thrombin factor that plays a role in successful feeding of the horn fly, Haematobia irritans. It has been isolated and characterized from saliva, and polymorphisms in the gene coding sequence have been previously reported. In the present study, the thrombostasin gene was analyzed from 60, field-collected flies from Camp Stanley, Texas and the allele and genotype frequencies were compared with previously published data for an Alabama field collection and a Texas in vitro colony-reared collection. Significant differences in genotype frequency and extent of genotypic diversity observed between the Alabama and Camp Stanley field collections may be attributable to host genetic differences. In addition, bull calves that were phenotyped as either high- or low-carriers were parasitized by horn flies that displayed a significantly different genotype distribution, supporting a possible explanation for horn fly host selection behavior, as evidenced by thrombostasin sequence analysis of 95 additional horn flies collected from respective hosts.     

Development of treated targets for controlling stable flies (Diptera: Muscidae)

Electrocution techniques were used to determine if treated targets similar to those used for tsetse control could be developed for stable fly control. In a series of two experiments, a half blue and half black (UK) 1 m2 target constructed of trigger cotton poplin was determined to be acceptable for development studies. In the first experiment, an average of 350 stable flies per hour (maximum 794 flies in 1 h) was collected using the UK target. A time-delayed circuit trial using untreated UK targets demonstrated that stable flies remained on or around the targets for at least 30 s. Two experiments were conducted with time-delayed circuits and UK targets treated with 0.1% lambda-cyhalothrin. In the first experiment, the number of flies collected using the 30 s on/off treated target treatment was not different from the number of flies collected using the other treatments. In the second experiment, the number of flies collected using the 30 s on/off treated target treatment was not different than the untreated target continuous or 30 s on/off treatments, but was significantly lower than the treated target continuous treatment. The number of flies collected with UK trigger targets was significantly higher than that for alsynite cylinder traps in two experiments. The mean number of flies collected during 22 1h assays using targets was 6.1-fold higher than that for alsynite traps, and the mean number of flies collected during 40 3 h using the targets also was 6.1-fold higher than that for alsynite traps. The results of this study indicate that treated cloth targets may be a viable addition for stable fly control programs.     

Influence of Superovulatory Protocols on In Vitro Production of Nellore (Bos indicus) Embryos

There are indications in the literature that delaying the period between ovarian superestimulation and ovum pick up (OPU) would induce follicles to a condition of initial atresia, which could be beneficial to oocyte development. In this work, we compared three protocols for OPU and in vitro production (IVP) of embryos, in Nellore cattle. Nellore cows (n = 18) were randomly allocated in three groups: Group 1 (OPU), Group 2 [Follicle stimulating hormone (FSH) and OPU] and Group 3 (FSH deprivation and OPU). Three OPUs were performed, and the animals were switched to a different group each time (crossover), in such a way that at the end of the experiment all cows received the 3 protocols. At random stage of the oestrous cycle (D‐2), all follicles ≥ 6 mm were aspirated to induce a new follicular wave 2 days afterwards (D0). In Group 1, OPU was performed on D2 and oocytes were processed to IVP. In Group 2, starting on D0, cows were superstimulated (FSH, Folltropin^®, 30 mg administered daily, i.m., during three consecutive days, total dose = 180 mg), and 6 h after the last FSH dose, they received exogenous luteinizing hormone (LH) (12.5 mg, i.m., Lutropin^®, D3). The OPU was performed 6 h after LH administration, i.e. 12 h after the last dose of FSH. Animals in Group 3 received the same treatment as those in Group 2, except that LH was administered 42 h after the last dose of FSH, and OPU occurred 6 h later. Therefore, in this group, follicles were deprived of FSH at 48 h. Both cleavage and blastocyst rates were similar (p > 0.05, anova ) among oocytes from Groups 1, 2 and 3, respectively: 77.4% (144/185) and 42.70% (79/185); 75.54% (105/139) and 31.65% (44/139); 63.52% (101/159) and 33.33% (53/159). However, hatched blastocyst rate was higher (p < 0.01) in Group 1 (30.27%, 56/185) when compared with Group 2 (11.51%, 16/139) or 3 (15.72%, 25/159). It is concluded that, contrary to previous work on European breeds (Bos taurus), ovarian superstimulation associated with deprivation of FSH and OPU (Group 3) did not increase IVP of Nellore embryos (Bos indicus). On the contrary, the highest hatched blastocyst rates were observed in oocytes from non‐superstimulated cows.     

Evaluation of a yearly insecticidal ear tag rotation for control of pyrethroid-resistant horn flies (Diptera: Muscidae).

From 1991 to 1997, the yearly alternated use of synergized pyrethroid (lambda-cyhalothrin + piperonyl butoxide) and organophosphate (pirimiphos-methyl) ear tags was evaluated for the control of two pyrethroid-resistant horn fly populations in Louisiana. At each site, weekly fly counts were used to assess product efficacy. Control achieved by synergized pyrethroid ear tag treatments was reduced from 7 to 2 weeks and from 4 to 0 weeks at St. Joseph and Winnsboro, respectively. Control by organophosphate ear tags decreased from 15 to 3 weeks and from 10 to 7 weeks at St. Joseph and Winnsboro, respectively. The rotation of synergized lambda-cyhalothrin and pirimiphos-methyl ear tags did not improve pyrethroid ear tag efficacy or prevent further development of resistance to the pyrethroid or OP compound.     

Differences in copper and selenium metabolism between Angus (Bos taurus) and Brahman (Bos indicus) cattle

A 2-yr study was conducted at the Range Cattle Research and Education Center, University of Florida – Institute of Food and Agricultural Sciences (IFAS) (Ona, FL), to evaluate differences in the metabolism of Cu and Se of Angus (Bos taurus) and Brahman (Bos indicus) cattle. Thirty-two pregnant beef cows (n = 8 Brahman and 8 Angus/yr) were enrolled in the study in the first trimester of gestation. This study consisted of three phases: 1) restriction (day 0 to 90), 2) supplementation (day 91 to 150), and 3) calving. During all three phases, cows were individually fed and housed in partially covered drylot pens. During the restriction and supplementation phases, cows were provided a 1.5 kg/d of a grain-based concentrate supplement, which was fortified with flowers of S (50 g of supplemental S/cow daily; restriction phase) or Cu and Se (100 and 3 mg/d of Cu and Se, respectively; supplementation phase). Blood and liver samples were collected from all cows at 30 d intervals and from both cows and calves within 24 h of calving. Colostrum and milk samples were collected at calving and 7 d after birth. All data were analyzed using the MIXED procedure of SAS, where cow and calf were the experimental unit. During the restriction phase, a breed × day effect (P = 0.03) was observed where Brahman had greater liver Cu concentration than Angus cows in all sampling days. For liver Se concentration, a tendency (P = 0.07) for a breed effect was observed where Angus cows tended to have greater liver Se concentration than Brahman. During the supplementation phase, breed (P < 0.001) and day (P < 0.01) effects were observed, where Brahman cows had greater liver Cu concentration than Angus. For liver Se concentration, a day effect (P < 0.001) was observed, where liver Se concentration increased (P < 0.001) from day 90 to 120 and remained unchanged (P = 0.86) until day 150. At calving, no effects of breed (P = 0.34) were observed for liver Cu concentration of cows; however, Brahman calves tended (P = 0.09) to have greater liver Cu concentration than Angus calves. For Se liver concentration at calving, Angus cows tended (P = 0.07) to have greater liver Se concentration than Brahman cows; however, no breed differences (P = 0.70) were observed for liver Se concentration of calves at birth. In summary, substantial differences in multiple indicators of Cu and Se status were observed between Angus and Brahman cattle, implying that Angus and Brahman cattle possibly have different mechanisms to maintain adequate Cu and Se status.