Ingestion of microfilariae from different sources by Culicoides nubeculosus (Diptera: Ceratopogonidae) through an artificial membrane

Laboratory-hatched Culicoides nubeculosus midges were membrane-fed on cattle blood containing various Onchocerca microfilariae which had been isolated from the hides of red deer (Cervus elaphus) and from Australian and native German cattle. All microfilariae were ingested through the membrane except those of O. tarsicola, which were too large. Onchocerca gibsoni and O. lienalis reached the thoracic musculature, but no further development occurred. Microfilariae of O. flexuosa developed to the "sausage" stage. Microfilariae of O. gutturosa developed to the infective larval stage within 9 days at a temperature of 25 degrees C.     

Distribution of microfilariae of Onchocerca lienalis and Onchocerca gutturosa in the skin of cattle in Germany and their development in Simulium ornatum and Culicoides nubeculosus following artificial infestation

Onchocerca microfilariae were isolated form the umbilicus and neck of 438 cow hides at the abattoir in Tübingen, F.R.G. The overall Onchocerca infection rate was 40.4%. The presence of Onchocerca lienalis and O. gutturosa microfilariae, which are difficult to distinguish by morphological criteria, was retrospectively demonstrated after artifically infesting Simulium ornatum and Culicoides nubeculosus and identifying the infective larvae recovered. Nine of 16 samples of umbilical microfilariae fed to C. nubeculosus through a latex membrane developed to O. gutturosa third stage larvae (L3). Six of seven umbilical samples injected into the thorax of S. ornatum yielded O. lienalis L3. In six infestation trials in which microfilariae were introduced both into S. ornatum and C. nubeculosus, O. lienalis L3 were recovered exclusively from simuliids, while O. gutturosa L3 developed only in midges. Of six umbilical skins tested by cross-infestation, one contained exclusively O. gutturosa microfilariae, four only O. lienalis microfilariae and one was infected with both species. Developmental success of O. lienalis microfilariae to L3 in S. ornatum following intrathoracic injection was 22% of the mean inoculum. O. gutturosa microfilariae, ingested by C. nubeculosus through a latex membrane, developed to L3 at a rate of 2.3% of the mean microfilarial uptake.     

Efficacy of ivermectin in paste and injectable formulations against microfilariae of Onchocerca cervicalis and resolution of associated dermatitis in horses

The efficacy of a single dose of ivermectin (0.2 mg/kg), in injectable or paste formulations, against microfilariae of Onchocerca cervicalis and associated skin lesions was evaluated in 20 naturally infected horses during midsummer months in Louisiana. All horses had clinical signs of dermatitis of the ventral midline and/or limbs, shoulders, thorax, and withers. Efficacy was monitored at 21, 42, and 63 days after treatment. Procedures done at these intervals included microfilarial counts of 6-mm skin biopsy specimens of affected ventral midline, grading of gross lesions, and photography and histologic assessment of ventral midline biopsy specimens. Microfilarial numbers were reduced to 0 by 21 days after treatment in all but one horse. Active lesions improved or were resolved completely by 63 days after treatment. Total inflammation, as judged by histologic methods, was reduced in all horses by 63 days after treatment, but there was a residual population of inflammatory cells in all horses. Adverse reactions after treatment were not observed in any of the horses.     

Efficacy of combined chemotherapy against gastrointestinal nematodes and Fasciola hepatica in cattle

Direct blood smear examination (using 0.05 ml of whole blood) detected 168 (80.9%) of 204 microfilaremic canine blood samples as determined by the modified Knott test for microfilariae (mff) of Dirofilaria immitis (using 1 ml of whole blood). Direct smear examination detected all of 134 microfilaremias greater than 50 mff ml(-1), but only 31 of 70 (44.3%) microfilaremias having less than 50 mff ml(-1). In a separate retrospective query of a database of 963 dogs with necropsy-confirmed heartworm infections, 834 (86.6%) were positive by the DiroCHEK heartworm antigen test, and 504 (52.3%) were microfilaremic by the modified Knott test. Only 2 (0.4%) of the microfilaremic dogs were DiroCHEK negative and another 18 (3.6%) were very weak positives. Although these microfilaremic dogs were not tested by direct smear, only one of the two DiroCHEK-negative and six of 18 weakly DiroCHEK-positive dogs had microfilaremias so low that a direct smear may have given a false negative result. Significant adverse reactions to either diethylcarbamazine or the macrolide endectocides have not been reported for microfilaremias less than 500 mff ml(-1), thus substitution of the direct smear for a concentration test for mff, such as the modified Knott test or membrane filtration, does not appear to increase the risk of an unexpected adverse reaction to heartworm prophylactic drugs. Such a substitution results in only a very slight decrease (on the order of 0.1%) in the overall sensitivity of heartworm screening, provided a test for mff is run concurrently with an antigen test. If a test for mff is the only screening test used, then substitution of a direct smear for a concentration test may decrease the sensitivity of heartworm screening by nearly 20%, depending on the prevalence of low level microfilaremias in the population of dogs tested.     

Variation in the level of grain defect light flecks and spots on cattle hides.

The occurrence of hide damage light flecks and spots was determined on tanned hides from 28 herds during a period of 8 to 12 months. Light flecks and spots are described as small areas of grain loss up to 3 mm in diameter that are seen on dyed crust cattle leather. Damage was found on 75.8% of all hides. The neck and shoulders were the anatomical region with the highest prevalence of damage. Sixty-eight per cent of all hides had light flecks and spots in this region. The forelimbs and dewlap were the anatomical region with the second highest occurrence with a prevalence of 39.1%. This distribution corresponded to the known distribution of lice in cattle. No significant differences were observed in age, sex, prevalence of lice in the herd assessed in March or infestations with different lice species. The frequency of light flecks and spots varied significantly during the year. The frequency was highest in the late winter and early spring, decreased significantly during the summer and was lowest in the autumn. This variation supported the importance of lice in the development of light flecks and spots and suggested a relatively long healing period for the damages induced by lice.     

A longitudinal study of a natural lice infestation in growing cattle over two winter periods

A group of 61 cattle which were naturally infested with lice was followed over two winter periods. Data were collected on the number of lice found at various body sites over this period. Summary statistics were produced and due to the repeated measured and hierarchical structure of the data, multi-level analysis was used to model the population dynamics of Bovicola bovis and assess the influence of the various hierarchical levels. A four level Poisson model was produced--level one, the individual number of lice at each parting; level two, the body site examined (shoulder, midline or rump); level three, the time of the inspection and level four, the animal. Seasonal fluctuation in lice numbers was modelled using a cosine function transformation of time. A seasonal pattern was seen in both year's with lice counts higher in the first than the second year. The midline area was the most sensitive to detection of B. bovis. Variance contributed by the individual animal was less than that contributed by the body site examined and the time of the inspection. The model suggested that lice numbers within the shoulder and rump sites were near random following a Poisson distribution, but aggregation of lice occurred at the midline site with the distribution between animals following a Negative Binomial Pattern. The midline site was the most sensitive site for detecting B. bovis. Infestation numbers were higher in the first winter when cattle were younger. In the second winter, there was no difference in lice numbers between cattle exposed to infection for the first or second time.     

A longitudinal study of a natural lice infestation in growing cattle over two winter periods

A group of 61 cattle which were naturally infested with lice was followed over two winter periods. Data were collected on the number of lice found at various body sites over this period. Summary statistics were produced and due to the repeated measured and hierarchical structure of the data, multi-level analysis was used to model the population dynamics of Bovicola bovis and assess the influence of the various hierarchical levels. A four level Poisson model was produced-level one, the individual number of lice at each parting; level two, the body site examined (shoulder, midline or rump); level three, the time of the inspection and level four, the animal. Seasonal fluctuation in lice numbers was modelled using a cosine function transformation of time. A seasonal pattern was seen in both years with lice counts higher in the first than the second year. The midline area was the most sensitive to detection of B. bovis. Variance contributed by the individual animal was less than that contributed by the body site examined and the time of the inspection. The model suggested that lice numbers within the shoulder and rump sites were near random following a Poisson distribution, but aggregation of lice occurred at the midline site with the distribution between animals following a Negative Binomial Pattern. The midline site was the most sensitive site for detecting B. bovis. Infestation numbers were higher in the first winter when cattle were younger. In the second winter, there was no difference in lice numbers between cattle exposed to infection for the first or second time.     

Onchocerca cervicalis in Horses from Southern Brazil

The objective of this study was to determine the occurrence of Onchocerca cervicalis in 1200 adult horses from rural areas of Rio Grande do Sul, Brazil. Umbilical skin specimens measuring 2 x 2 cm were minced, suspended in 10 ml of distilled water and incubated at room temperature overnight. The liquid volume was centrifuged and the sediment was screened for microfilariae. The ligamentum nuchae were totally removed, preserved in ice and dissected for the detection of adult forms of Onchocerca cervicalis. Microfilariae of Onchocerca cervicalis were detected in midventral skin biopsy samples in 215 (17.9%) of 1200 horses examined and the adult worms were recovered from 200 (16.6%) ligamentum nuchae from the same animals. These findings provide a basis for further studies to determine infection rates in horses from other regions of the state and to identify the intermediate host that transmits the disease.     

Clinical evaluation of a right-sided prophylactic gastropexy via a grid approach

A new technique is described for a prophylactic incisional gastropexy via a right-sided grid approach (i.e., minilaparotomy). A pilot study showed comparable tensile strength between a traditional ventral midline approach and the grid approach. Six client-owned dogs were selected for the procedure. Four weeks postoperatively, a barium gastrogram was performed to assess stomach/gastropexy position. Complications included a seroma and postoperative discomfort. This technique should be considered for any at-risk breed that is not overweight. The right-sided grid approach to a prophylactic gastropexy was less invasive than a ventral midline approach and resulted in a stable gastropexy at 4 weeks postoperatively in five of six dogs.     

Efficacy of ivermectin against Onchocerca cervicalis microfilarial dermatitis in horses

Forty horses having microfilariae of Onchocerca cervicalis in association with dermatitis, alopecia, and pruritus on the ventral midline were given a single IM injection of 0.2 mg of ivermectin/kg of body weight (June to August 1981). Microfilarial counts in the 40 horses ranged from 18 to 42,446 microfilariae/skin snip on the day of treatment, and histopathologic examination of these skin sections indicated a chronic eosinophilic dermatitis. Numerous microfilariae were in the dermis, but there was no consistent relationship between the presence of microfilariae and the severity of the inflammatory reaction. In all 40 horses, skin snips taken 4 to 33 days after treatment were negative for microfilariae, and a marked clinical improvement occurred 2 to 3 weeks after treatment, when the lesion was replaced by healthy skin and new hair. Twenty-four hours after treatment, an edematous reaction occurred on the lower portion of the abdomen of 4 (10%) horses and within the area of the lesion in 6 (15% horses). The reactions disappeared within 24 to 72 hours, irrespective of whether horses were treated with corticosteroids. When further skin samples were taken from 15 horses 4 to 9 months later, 9 of them were free of microfilariae and the 6 others had only low counts (17 to 97). An additional 7 infected horses, treated when there was little chance of reinfection (November to December 1981), were all free of microfilariae at 6 to 10 days and 4 to 5 months after treatment. The marked clinical improvement in all horses after disappearance of microfilariae from the dermis indicates that microfilariae are involved in the cause and pathogenesis of the dermatitis.     

Occurrence,Antibiotic Resistance and Molecular Characterization of Listeria monocytogenes in the Beef Chain in the Republic of Ireland

This study investigated the occurrence, concentration and key characteristics of Listeria monocytogenes in beef chain samples (n = 1100) over a 2‐year period (July 2007–June 2009). Listeria monocytogenes was isolated from bovine hides (27%), pre‐chill carcasses (14%) and ground beef (29%), but not from ready‐to‐eat (RTE) beef. The concentration of the pathogen in the majority (95%) of contaminated samples was low and detected by enrichment only. The highest concentrations recovered (100–200 CFU/g) were in ground beef samples. The most commonly isolated serotype group was 1/2a (58%) followed by 4b (12%), 1/2b (10%) and 1/2c (6%). A small portion (<5%) isolates had demonstrated resistance to key anti‐microbials including ampicillin, vancomycin and gentamycin which are recommended treatment options for listeriosis. Pulsed‐field gel electrophoresis showed indistinguishable profiles for a number of isolates recovered from the hide and carcass (after slaughter and dressing) of the same animals, highlighting the role of hides as a source of contamination. Equally, indistinguishable pulsotypes for isolates recovered at different stages and time points (up to 6 months apart) in the beef chain demonstrated the persistence of specific clones in the factory, process and distribution environments. Overall, the study demonstrated a high prevalence of clinically significant L. monocytogenes entering and progressing along the beef chain and highlights the needs to control cross‐contamination during beef processing and distribution and the need for thorough cooking of raw beef products.     

The effect of eradication of lice on the occurrence of the grain defect light flecks and spots on cattle hides.

The influence of an eradication programme for lice on the prevalence of light flecks and spots on cattle hides was studied in 33 dairy cattle herds during a period of two and a half years. Lice were eradicated from the main group of herds after 9 to 12 months and the quality of the hides before and after treatment was compared. Hides from slaughtered animals were collected during the study period, tanned and examined with special emphasis on the occurrence of the grain damage light flecks and spots. The prevalence of hides without light flecks and spots increased from 24.2% before treatment to 61.6% after treatment. The prevalence of hides free from the damage increased significantly in all examined anatomical regions. The improvement in hide quality was most marked in the shoulders and neck region which corresponded to the major predilection site of cattle lice. The prevalence of hides with light flecks and spots started to decrease in the first period (2-40 days) after eradication. The changes after treatment suggested that most healing process took place over a period of about 4 months. The eradication programme eliminated the seasonal variation in the prevalence of light flecks and spots which was present before treatment.     

The effect of climate on the presence of Culicoides imicola in Italy

A model was developed to classify the Italian territories in relation to their suitability to harbour populations of Culicoides imicola and, as a consequence, also able to sustain a bluetongue (BT) epidemic. Italy was subdivided into 3507 10 x 10 km cells. In 546 cells at least one collection was made. The cell was considered the unit for all subsequent analyses. Culicoides were collected using Onderstepoort-type blacklight traps. Some traps were operated weekly at chosen sites; the remainder were moved almost daily to new sites. Only the results obtained during the peak August-November period were used, to exclude bias caused by the seasonality of C. imicola. Climate data for the period 1999-2001 were obtained from 80 weather stations. Multiple logistic regression was performed using the presence or absence of C. imicola in a specific cell as the dependent variable. Annual means of daily values for minimum temperature and minimum relative humidity, and the mean altitude above sea level, were the independent variables. The probability of occurrence of C. imicola in each grid cell was used to create a prediction map for Italy. The model was able to correctly classify 77.5% of the 546 grid cells in which at least one collection had been made. Culicoides imicola was found frequently through much of Sardinia, in parts of southern Italy, and further north along the Tyrrhenian coast, but was absent from along most of the Adriatic coast, and the internal mainland, and from most of Sicily. Six detailed maps are provided. Also mapped are areas where the probability of the occurrence of C. imicola is lower than 5%. This identification of possible mountainous C. imicola-free areas in central Italy could facilitate safer animal trade and transhumance, even if BT infections in traded animals or moving stock, were to go undetected. Needless to say this depends upon no cool-adapted species of Culicoides being involved in the transmission of BT disease.     

Evidence of Onchocerca fasciata (Filarioidea: Onchocercidae) in camels (Camelus dromedarius): I-prevalence, nodular lesions appearance and parasite morphology

Examination of 3376 imported and 200 local Egyptian camels was carried out during the period extending between September 1997 and August, 1998. These animals were carefully examined for the presence of Onchocerca fasciata nodules and subsequently for O. fasciata adult parasites. Results of this study revealed that imported camels had the higher infection rate (2.75%), while those of local origin showed no palpable or detected Onchocerca nodules. Distribution of these nodules was mainly on the two sides of abdomen, hind limbs (concentrated in thigh region) and fore limbs particularly on the shoulders and nuchal ligament. This distribution varied according to the degree of infection. Searches for microfilariae were also performed using either blood samples, or in the subcutis and fascial sheath near or around the detected nodules.     

Bovine and equine onchocerciasis in eastern North America with a discussion on cuticular morphology of Onchocerca spp. in cattle.

Skin sections and/or the ligamentum nuchae and ligamentum gastrolienale were examined from twelve bovine carcasses obtained from southern and eastern Ontario and from Quebec. Of these, seven were shown to be infected with Onchocerca gutturosa and/or Onchocerca lienalis. The morphology of the adult female cuticle is discussed. Skin sections from 43 equine carcasses from a slaughter house in Grenville, Quebec were examined and microfilariae of Onchocerca sp. were recovered from 32 (74%). There are probably referable to Onchocerca cervicalis.     

Molecular genetic comparison of Onchocerca sp. infecting dogs in Europe with other spirurid nematodes including Onchocerca lienalis

In the past 15 years, subconjunctival onchocercosis has been reported from 63 dogs in south-western United States (Arizona, California, Utah) and Southern and Central Europe (Germany, Greece, Hungary, Portugal, Switzerland). To reveal the taxonomic status of the parasite responsible for these infections, fragments of the mitochondrial cytochrome oxidase subunit I (COI) and NADH dehydrogenase subunit 5 (ND5) genes of three European strains of canine Onchocerca sp. and the 16S ribosomal RNA (16S rRNA) gene of their Wolbachia endosymbionts were sequenced and compared to the homologous sequences of other spirurid nematodes. The evolutionary divergence between COI and ND5 gene sequences of Greek, Hungarian and Portuguese strains of canine Onchocerca sp. were similar in magnitude to that seen within Thelazia callipaeda or Onchocerca lienalis. The evolutionary divergence between the sequences of canine Onchocerca sp. and other Onchocerca spp. including O. lienalis were similar or higher in magnitude to that seen between other Onchocerca spp. The results of the current and earlier phylogenetic analyses indicate that canine Onchocerca sp. separated from other Onchocerca spp. early in the evolution. Based on the similar clinical pictures, the identical morphology of nematodes and the sequence analyses of COI and ND5 genes of the worms and 16S rRNA gene of their wolbachiae, the Onchocerca worms isolated from European dogs appear to belong to the same species. The results support the earlier biological and morphological arguments that a distinct species, most likely O. lupi originally described from the subconjunctival tissues of a Caucasian wolf is responsible for canine ocular onchocercosis in Europe.     

Teat onchocercosis in cows with reference to prevalence, species involved and pathology

The present study reports teat onchocercosis in cows in the province of Kars in north eastern Turkey with reference to the prevalence, species involved and pathological findings. In the study, 600 cows of various breeds and ages slaughtered in the local abattoir were examined and 145 (24%) were found to have various lesions on their teats, of which 45 (31%) were infected with three Onchocerca species (Onchocerca gutturosa, O. lienalis and an unidentified Onchocerca spp.), identified according to their morphological characteristics. Gross pathological examination of the teats with lesions revealed scabies and chaps of varying sizes, healed sores and small nodules. Histopathologically, large numbers of microfilariae were commonly observed within the collagenous fibre bundles of the dermis, with only a few microfilariae in the dermis in some cases. The microfilariae were also often found to have accumulated around the perivascular spaces and were frequently associated with infiltration by inflammatory cells, predominantly eosinophil leukocytes and mononuclear cells. In two cases, microfilariae were discovered exclusively in the lumen of the capillaries of the dermal papillaries. In one case, in which microfilarae were not encountered, sections of adult worms surrounded by neutrophil leukocytes were detected free in the teat canal. The presence of extensive teat lesions along with microfilariae of the Onchocerca species as found in this study warrants further assessment of the impact of onchocercosis on dairy farming.     

Ultrasound-assisted collection of cerebrospinal fluid from the lumbosacral space in equids

OBJECTIVE: To describe ultrasonographic landmarks for use in collection of CSF from the lumbosacral region in equids. DESIGN: Prospective study. ANIMALS: 37 equids (27 with neurologic disease and 10 with nonneurologic disease). PROCEDURES: Standing equids (n = 17) were sedated with detomidine hydrochloride (0.006 to 0.01 mg/kg [0.003 to 0.005 mg/lb], IV) followed by butorphanol tartrate (0.01 mg/kg, IV) and restrained with a nose twitch for collection of CSF. The CSF was collected from 20 laterally recumbent equids (10 sedated and 10 immediately after euthanasia). Anatomic landmarks were identified ultrasonographically. Height at the dorsal point of the shoulders, body weight, depth of the spinal needle, number of attempts to collect CSF, and cytologic evaluation of CSF were recorded. RESULTS: Lumbosacral puncture cranial to the cranial border of the most superficial location of both tuber sacrale along the midline was consistently successful for CSF collection (35/37 equids). Two horses had anatomic abnormalities that precluded CSF collection. Mean number of attempts to collect CSF per animal was 1.1. Height and body weight were strongly correlated with needle depth for CSF collection. Pelvic and sacral displacement was observed in several laterally recumbent animals, which resulted in discrepancies of the midline between the cranial and caudal aspects of the vertebral column. In most equids, the spinal needle was aligned on the midline of the caudal aspect of the vertebral column. CONCLUSIONS AND CLINICAL RELEVANCE: Ultrasonography was a useful aid for collection of CSF from the lumbosacral space and decreased the risk of repeated trauma and contamination in equids.     

驴皮低温酶解脱毛方法及其对阿胶质量的影响

[目的]开发一种适合阿胶产业化生产的高效驴皮脱毛方法。[方法]低温条件下（4 ℃）,选用1%碱性蛋白酶和1%中性蛋白酶对醒发的干驴皮开展酶解脱毛试验,比较2种酶解方法处理下驴皮的脱毛时间、脱毛率,并对脱毛过程中驴皮的气味进行评价。以1%碱性蛋白酶和1%中性蛋白酶酶解处理驴皮以及机械脱毛鲜驴皮为材料熬制阿胶,比较不同原料熬制阿胶的出胶量、出胶率、外观、气味等;测定以2种蛋白酶酶解处理的驴皮为原料熬制的阿胶质量指标,包括水分、灰分、水不溶物以及4种氨基酸含量,并与《中国药典》2015版一部中提供的相应质量标准进行对比。[结果]低温条件下（4 ℃）,利用1%碱性蛋白酶（pH值=10.5）或1%中性蛋白酶（pH值=7.5）酶解处理驴皮第2天表现出脱毛效果,处理第5天醒发的干驴皮脱毛率可达90%,有臭味产生但非驴皮腐败臭味,而是脱毛过程中产生的正常气味。以1%碱性蛋白酶、1%中性蛋白酶酶解脱毛后的驴皮以及机械脱毛鲜驴皮为原料熬制阿胶,出胶量分别为113.0、109.4、129.3 g,出胶率分别为13.8%、13.1%、13.2%;2种酶解处理驴皮熬制的阿胶色红、气味芳香,与鲜驴皮熬制的阿胶相比色泽更鲜亮。2种酶解处理驴皮熬制阿胶的水分、灰分、水不溶物均符合并优于《中国药典》2015版一部中的质量标准,L-羟脯氨酸、甘氨酸、丙氨酸、L-脯氨酸含量也远高于《中国药典》2015版一部中的质量标准。[结论]低温条件下（4 ℃）,1%碱性蛋白酶和1%中性蛋白酶可对醒发干驴皮进行有效酶解脱毛,且脱毛后驴皮熬制的阿胶质量符合《中国药典》2015版一部标准。