The epidemiology of Parafilaria bovicola in the Transvaal Bushveld of South Africa

A total of 20 375 flies collected off cattle on 12 farms over 36 months were identified and examined for 3rd stage P. bovicola. The 3 vector species accounted for 64.1% of the flies collected and were the only fly species found to be infected. Musca lusoria was clearly the dominant vector fly, although large numbers of Musca sp. A appeared regularly between February and April each year. This phenomenon, coupled with high numbers of M. lusoria throughout most of the year, led to an increase in the numbers of vector flies from their lowest level in June to a peak in February-April. Of the 13 070 vector flies examined for 3rd stage larvae only 64 (0.52%) were positive; of these 41 were M. lusoria and 17 Musca sp. A. No positive male flies were found. Incubation of wild-caught flies for up to 13 days at 27 degrees C noticeably increased the larval recovery rate. Flies were found to be infected mainly from August-March. Infected M. lusoria were recorded from July-March and infected Musca sp. A from January-May. Only 6 infected M. xanthomelas were collected and this was during the period August-December, when most ovipositional blood spots occur on cattle. It is concluded that P. bovicola transmission in the Bushveld is not correlated with peak periods of bleeding but rather with high numbers of vector flies, the various species augmenting each other so that transmission may take place almost throughout the year.     

The colonization and life-cycles of Musca lusoria, Musca xanthomelas and Musca nevilli, vectors of Parafilaria bovicola in South Africa

Thriving, permanent colonies of Musca xanthomelas and later of Musca nevilli were successfully established. However, because of the low reproduction potential of Musca lusoria a small colony only was kept for a limited period until life-cycle studies were completed. Larvae were reared on fresh dung from cattle fed lucerne, while in general adults were fed 0.3% citrated ox-blood, whole milk powder, sugar crystals, fresh dung and water. M. nevilli could be colonized only when ox-liver was substituted for ox-blood. A comparison of the life-cycles of M. lusoria and M. xanthomelas under laboratory conditions at a constant temperature of approximately 27 degrees C, 60% R.H. and 24 h illumination revealed major differences between these 2 vector species. M. lusoria deposits single larvae at intervals of approximately 2 days and a female can produce up to 27 in her life-time. An M. xanthomelas female can lay up to 4 batches of eggs, with as many as 33 eggs per batch, at intervals of approximately 5 days. A single female can produce a maximum of 94 eggs. M. lusoria, however, showed survival advantages over M. xanthomelas in that its larvae reached the pupal stage at least a day sooner and its adults survived more than twice as long. The life-cycles of M. xanthomelas and M. nevilli were similar in the laboratory, except for adult dietary requirements. The mean number of mature oocytes in the ovaries of M. nevilli, however, was only 15.7 compared with 26.1 in M. xanthomelas.     

The effect of arsenical dips on Parafilaria bovicola in artificially infected cattle in South Africa

The possible adverse effect of arsenical tick control dips on Parafilaria bovicola infections was investigated in 48 artificially infected cattle. A treatment group of 24 cattle was dipped in a plunge dip containing 1600 ppm arsenic trioxide. A control group of the same size was dipped in an organophosphate dip containing a mixture of chlorfenvinphos and dioxathion. Regular weekly to 3-weekly dipping had no effect initially on the prevalence of ovipositional blood spots of P. bovicola in either group. However, from 4 months after bleeding commenced there was a significant reduction in blood spots in the arsenic-dipped cattle and, on slaughter at 12-14 months after infection, the arsenic group had significantly fewer live worms and fewer carcass lesions. Arsenic residues in muscle samples of treated cattle were 11.6 times higher than in the controls. It is proposed that arsenic residues in the sub-cutaneous muscle layers increase with repeated dipping until a level toxic to P. bovicola is finally reached. Older cattle would therefore be refractory to infection and their carcasses at slaughter would not be affected.     

The longevity of adult Parafilaria bovicola and the persistence of their associated carcass lesions in cattle in South Africa

The slaughter of naturally-infected heifers and oxen at regular intervals after the first P. bovicola ovipositional blood spots appeared revealed that no female worms with embryonated eggs could be found after 259 days, no live worms after 372 days, and no carcass lesions after 519 days. In one bull, however, blood spots and carcass lesions persisted throughout 2 seasons. With the possible exception of bulls, therefore, annual reinfection of cattle appears to be necessary for the continuation of the transmission cycle of P. bovicola.     

Parafilaria bovicola (Tubangui 1934) established in Swedish cattle

Parafilaria bovicola was recorded in Swedish cattle for the first time in 1978. The parasite causes carcass lesions with a maximum incidence from February to July but already in the previous November the carcass lesions appear regularly in the form of yellow oedema. In February and onward the oedema may be extensive with a greenish-yellow colour. The lesions are most prominent in bulls and more or less negligible in cows. However, the cows are considered to be important reservoirs for the parasite from one season to the next. Bleedings are seen from March until August and the infective larvae are transmitted to the cattle from early June by the intermediate host Musca autumnalis. Parafilaria lesions look remarkably like those caused by contusions during handling and transport previous to slaughter. Eosinophile infiltrations can be regarded as a constant diagnostic feature of parafilariosis.     

The control of Parafilaria bovicola transmission in South Africa

Ivermectin treatment of all cattle on a badly infected farm failed to interrupt the transmission of P. bovicola, even though ovipositional blood spots were drastically reduced in numbers for an entire summer season following treatment. Regular weekly to fortnightly dipping of all cattle in 50 ppm deltamethrin immediately reduced vector fly numbers to less than 1 fly per cow face. Sustained dipping for 9 months effectively reduced P. bovicola transmission from approximately 50% to less than 2%. However, cessation of fly control led to a return to predipping P. bovicola infection levels. Ovipositional blood spot counts and the ELISA technique for evaluating P. bovicola infection in a herd were compared and were both effective methods. Best results for the blood spot method, however, are obtained in spring at the peak of the bleeding season whereas the ELISA method does not have this limitation.     

Rearing of Culicoides nubeculosus (Diptera: Ceratopogonidae) by natural or artificial feeding in the laboratory

A technique for the mass rearing of Culicoides nubeculosus in the laboratory is described. Female midges were fed either on fresh or deep-frozen, defibrinated cattle blood (-70 degrees C) through latex membranes, or on anesthetized white mice. Feeding rates of up to 90% were obtained on the latex membrane, whereas only 41% of the midges fed on mice. The best oviposition rates of greater than 50% were achieved after feeding either on the latex membrane with fresh cattle blood or on mice. An average of greater than 100 eggs per female were deposited. The highest larval hatching rate was observed after feeding with fresh blood; about half of the larvae developed to the adult stage. A reproduction index was defined for the colony based on the feeding rate, oviposition rate, larval hatching rate and development to the adult stage. The highest reproduction index was obtained when the midges were fed on fresh cattle blood through the membrane.     

Preparation and evaluation of the specificity of Parafilaria bovicola antigen for detection of specific antibodies by ELISA

An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies in bovine sera against Parafilaria bovicola nematodes was developed and its sensitivity was compared with the immunodiffusion (ID) method. An exoantigen of P. bovicola which was shown to contain four major polypeptides was used in these procedures. The serological reactivity of the antigen polypeptides was defined by using the enzyme-linked immunoelectrotransfer blot technique (EITB) and whole-worm extract proteins. It identified only four serologically reactive polypeptides with sera from one experimentally infected calf and a verified field case. These two positive sera reacted mainly with four major antigens which coincided in molecular weights of the polypeptides of the exoantigenic preparation, namely, 43, 39, 28 and 25 KDa. Calves experimentally infected with P. bovicola showed a positive reaction with ELISA at 4 months after inoculation, and after this period a rapid increase in serum antibody response occurred. In these cases the ID reaction was observed for the first time at 7 months after inoculation. The specificity of an ELISA method using crude exoantigen preparation of P. bovicola was tested for the diagnosis of bovine parafilariasis. No cross-reactivity was detected when the P. bovicola exoantigen preparation was tested against sera from calves experimentally infected with Onchocerca lienalis, as well as against the sera from cattle naturally infected with Dictyocaulus viviparus or from cattle chronically infected with Ostertagia ostertagi. In addition, testing of 740 field sera from cattle in areas non-endemic and endemic for P. bovicola indicated a specificity of the antigen preparation used. Forty sera from laboratory-confirmed field cases of P. bovicola infection were tested by ELISA and immunodiffusion. All of these sera were ELISA positive, whereas only 70% of these were positive in the ID test. Seven (2.1%) of 328 sera from 21 herds from non-endemic P. bovicola areas were ELISA positive, as opposed to none in the ID test. Of the 94 sera from six herds in areas endemic for P. bovicola infection, 51 (54%) were ELISA positive whereas only 24 (26%) were positive in the ID test. When 56 slaughtered cattle, with varying degrees of meat condemnations due to parafilariasis, were tested for P. bovicola specific antibody, 91% of the serum samples were positive by ELISA. These results suggest that the exoantigen of P. bovicola can be used in a sensitive and reliable serological detection of parafilariasis by ELISA.     

蛋白质水平对肉牛屠宰指标的影响

[目的]探讨不同蛋白质水平对肉牛屠宰指标的影响。[方法]选用5个营养水平的精料补充料配方,分别饲喂5个试验组肉牛,并与对照组进行比较。[结果]饲喂粗蛋白水平较高的精料补充料有利于胴体脂肪的沉积和眼肌面积的增大,有利于肉牛的胴体测量指标的提高。蛋白质水平对脏器重量的影响主要集中在头、皮、肝、肾、胆囊、膀胱和横膈膜,对消化器官重、心重、肺重、脾重的影响没有发现规律性。[结论]提高肉牛屠宰指标可以通过补充蛋白质来实现。     

Epidemiology of wildebeest-derived malignant catarrhal fever in South Africa: inability to transfer the disease with an African face fly Musca xanthomelas (Diptera: Muscidae)

Under experimental conditions an African face fly (Musca xanthomelas) preferred to feed on cattle dung when provided with a choice of 3 different meals namely sucrose, cattle dung and blood. Flies starved overnight fed well on the eyes of cattle and rabbits, but were reluctant to feed again within 2 h after being allowed to feed on cell culture medium or on the eyes of wildebeest, and when they did feed, they preferred to feed on the external side of the eyelids and on the coagulated material in the medial canthus of the eye. Under field conditions flies were rarely seen to feed on the eyes of immobilized wildebeest. Although M. xanthomelas became infected with Alcelaphinae herpes virus 1 (AHV-1) when they fed on infective wildebeest tears or cell culture medium, they lost the virus within 5 h, and recovery of infective AHV-1 particles from regurgitated cell culture medium was limited to the first 30 min after feeding. AHV-1 could not be transmitted by flies to cattle or rabbits. The failure to transfer the virus with flies can be ascribed to their reluctance to feed on cattle or rabbits shortly after they have consumed a protein rich meal, the rapid inactivation of ingested virus and the relatively high titre of virus necessary to infect cattle via the ocular route. Furthermore, it is believed that under natural conditions flies that have emerged from cattle dung will be inclined to stay with cattle where food is freely available.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization and purification of Parafilaria bovicola antigens by chromatofocusing to enhance specificity in serodiagnosis

A study was conducted to determine if the purification of Parafilaria bovicola antigens can increase the specificity of serodiagnosis of parafilariasis in enzyme-linked immunosorbent assay. Antigens released from adult worms of P. bovicola were separated by chromatofocusing on a polybuffer exchanger of the pH range 7.3-4.0 Polypeptide analysis by sodium dodecyl sulphate polyacrylamide gel electrophoresis showed the presence of four major polypeptides with MWs of 41, 36, 24 and 20 kDa. Additional biochemical characterization identified the 24- and 20-kDa polypeptides as hydrophobic glycoproteins. The chromatofocusing purification procedures were also applied for separation of a whole-worm extract. Again, the 41- and 36-kDa antigens were identified in separate peak fractions. Using ELISA, it was shown that the 41- and 35-kDa antigens were recognized by bovine antibodies specific for P. bovicola, but not by other sera collected from cattle infected by Onchocerca gutturosa, Onchocerca lienalis, Ostertagia ostertagi and Dictyocaulus viviparus. The serological evaluation strongly suggests that the 41- and 36-kDa antigens are P. bovicola specific.     

Studies on Parafilaria bovicola Tubangui, 1934, 2. Chemotherapy and pathology

The filaricidal effects of trichlorphon, arsenic trioxide, sodium antimony biscathecol disulphonate and nitroxynil against Parafilaria bovicola were the subject of this investigation. Levamisole hydrochooride was retested in a separate trial to assess the time required for healing after successful treatment. A comparison of carcass lesions and the percentage of lesion area in untreated controls with those in treated animals reconfirmed the efficacy of levamisole hydrochlorice, while nitroxynil gave more promising results. Both lesions and lesion area were reduced by 76% by the former and by 93% by the latter compound. The other drugs only slightly suppressed P. bovicola activity. After treatment with levamisole hydrochloride, active lesions were still present for 1-4 weeks and visible lesions on slaughter for 8 weeks. Since the visible lesions during the 8 weeks varied from acute to chronic and as they disappeared 9 weeks post treatment, it is suggested that provision should be made for a healing period of at least 9 weeks.     

Seasonal abundance and distribution of Parafilaria bovicola ovipositional blood spots on cattle in South Africa

More than 23 000 cattle of both sexes and different ages were examined for blood spots caused by egg-laying females of P. bovicola. Although these studies extended over four years and involved 5 farms in different parts of the Transvaal Bushveld, the overall results were the same. Ovipositional bleeding was strongly seasonal with blood spots first appearing in winter (June), reaching a peak in spring (September-November) and thereafter declining rapidly as summer progressed. In a single year at Zoutpan Research Station up to 92,1% of the 1st year heifers had already bled by November and this proportion increased only slightly to 95,1% by the end of the bleeding season (May). The number of blood spots per animal showed a similar seasonal abundance except for a second peak of abundance in June for 1st year heifers and oxen. The prevalence of blood spots in cattle of different ages and sex varied markedly. At Mara Research Station half as many oxen bled in their 2nd year as in their 1st year, while at Zoutpan 19,2% fewer heifers bled in their 2nd year than in their 1st. Bulls bled the most, then 1st year oxen, 1st and 2nd year heifers and 2nd year oxen, with breeding cows bleeding the least. A high female hormone level appears to be associated with the development of immunity. The shortest period from birth to 1st blood spot (the apparent prepatent period) was 191 days, while 81,8% of oxen bled for the 1st time within 279 days after birth.(ABSTRACT TRUNCATED AT 250 WORDS)     

Simulium jenningsi Malloch (Diptera: Simuliidae): a vector of Onchocerca lienalis Stiles (Nematoda: Filarioidea) in New York

Biting flies were collected from the umbilical areas of Onchocerca lienalis-infected cattle in New York (state) from June through September of 1980. Of the 766 flies collected, 705 were Simulium jenningsi. Microfilariae were detected in the midguts of 37 (50%) of 73 females dissected immediately after the flies had fed. The mean number of larvae per positive fly (fly with microfilariae) was 15.2. Third-stage larvae were recovered from 25 (21.9%) of 114 S jenningsi dissected 8 to 13 days after they had fed on the infected cattle; the mean number of 3rd-stage larvae per positive fly was 3.5. Dissections of flies performed on days 1 through 7 after feeding yielded various numbers of 1st and 2nd-stage larvae from the thoracic muscles. Ovarian dissections performed on 304 S jenningsi attacking cattle indicated an overall parous rate of 58%. Naturally occurring infections with filarial larvae indistinguishable from O lienalis were found in 7.3% of the parous females. Three of these flies, or 1.7% of the parous collection, harbored 3rd-stage larvae. The onset of naturally occurring filarial infections in the population of S jenningsi coincided with a peak in the parous rate in late June. Thereafter, filarial infections were generally detected when the parous rate was above 50%.     

Role of insects in the transmission of bovine leukosis virus: potential for transmission by stable flies, horn flies, and tabanids

The ability of stable flies (Stomoxys calcitrans), horn flies (Haematobia irritans), and tabanids (Diptera: Tabanidae) to transmit bovine leukosis virus (BLV) was investigated. Stable flies and horn flies were fed on blood collected from an infected cow, and the flies' mouthparts were immediately removed, placed in RPMI-1640 medium, ground, and inoculated into sheep and calves. Infection of sheep occurred with mouthparts from as few as 25 stable flies or 25 horn flies. However, sheep were not infected when removal of stable fly mouthparts was delayed greater than or equal to 1 hour after blood feeding. Infection of calves occurred after inoculation of mouthparts removed immediately after feeding from as few as 50 stable flies or 100 horn flies. Infected blood, applied by capillary action to the mouthparts (labella) of 15 deer flies (Chrysops sp) and a single horse fly (Tabanus atratus) caused infection in each of 2 sheep. Infection did not occur in 2 calves inoculated daily for 5 days with mouthparts from 50 horn flies collected after feeding on a BLV-infected steer. Four calves receiving bites from 75 stable flies interrupted from feeding on a BLV-positive cow also were not infected. Seronegative cattle held for 1 to 4 months in a screened enclosure with positive cattle in the presence of biting flies were not infected with BLV. The feeding behavior of each insect is discussed to assess their potential as vectors of BLV.     

The detection of lumpy skin disease virus in samples of experimentally infected cattle using different diagnostic techniques

Lumpy skin disease (LSD) is a disease of cattle, primarily in Africa and Madagascar and rarely in the Middle East. It is caused by a capripoxvirus that belongs to the family Poxviridae. The disease is of economic importance in endemic areas. Effective control of LSD requires accurate and rapid laboratory techniques to confirm a tentative clinical diagnosis. Comparative studies on different diagnostic tests used at different stages of the disease have not been done. The aim of this study was to compare several of these tests. Six seronegative bulls, between 11 and 20 months of age, were infected intravenously and kept in an insect-free facility. The course of the infection was monitored. During a 3-month period blood samples and skin biopsies were collected for virus isolation and polymerase chain reaction (PCR). Skin biopsies were also examined using transmission electron microscopy (TEM). The incubation period in infected animals varied from 4-5 days. The length of the viraemic period did not correlate with the severity of clinical disease. Viraemia was detected from 1-12 days using virus isolation and from 4-11 days using the PCR, which is longer than has previously been reported. Virus was isolated from skin biopsies until Day 39 post infection (p.i.) and PCR could demonstrate viral DNA until Day 92 p.i. Transmission electron microscopy of negatively stained skin biopsies detected LSD virus only in one of the four bulls that developed skin lesions until Day 33 p.i. The PCR was a fast and sensitive method to demonstrate viral DNA in blood and skin samples. It could detect viral nucleic acid in skin lesions 53 days longer than virus isolation. Virus isolation from blood and skin samples was sensitive and reliable, but as a single test it may be too time-consuming to use although this depends on how rapidly the diagnosis must be confirmed. In conclusion, this study showed the PCR to be superior in detecting LSD virus from blood and skin samples. However, virus isolation is still required when the infectivity of the LSD virus is to be determined.     

San Miguel sea lion virus fed to mink and pigs.

Mink became infected with San Miguel sea lion virus when fed ground meat from seal carcasses showing vesicular-like lesions in the skin. The mink also contracted the infection when they were fed San Miguel sea lion virus infected pig meat or cell culture propagated virus. San Miguel sea lion virus infection in mink was inapparent but the virus was isolated from blood and rectal swabs. Pigs treated similarly with the same virus preparations given to mink developed a severe vesicular disease syndrome similar to that produced by vesicular exanthema of swine. In a separate trial, pigs fed a large sample of commercial ground seal meat did not develop disease signs or antibodies. Further work is needed to assess the hazard of introducing San Miguel sea lion virus into swine on the same premises when potentially San Miguel sea lion virus infective seal meat is fed to mink.     

Infection of sheep and goats with bovid herpesvirus 2

Sheep and goats were shown to be susceptible to experimental infection with bovid herpesvirus 2 (BHV2), administered by either the intradermal or intravenous route. Lesions developing in sheep following intradermal inoculation of virus were similar to those in cattle inoculated intradermally, whereas the lesions in goats resolved without ulceration or scabbing. Disseminated circumscribed skin lesions developed in sheep and goats given BHV2 by the intravenous route. These lesions resolved in four to eight days without significant effect on the skin. BHV2 was isolated from skin lesions of sheep, goats and cattle that were infected intravenously, from sheep and cattle infected intradermally and from the leucocytes of the three species following intravenous inoculation of virus. Latent infection of sheep and goats was demonstrated following corticosteroid treatment 60 days after infection.     

The application of PCR-ELISA to the detection of Trypanosoma brucei and T. vivax infections in livestock

Teneral tsetse flies infected with either Trypanosoma brucei or T. vivax were fed on healthy cattle. Blood samples collected daily from the cattle were examined by microscopy for the presence of trypanosomes, in thick smear, thin smear and in the buffy coat (BC). All the cattle fed upon by infected tsetse developed a fluctuating parasitaemia. DNA was extracted from the blood of these cattle and subjected to polymerase chain reaction (PCR) using oligonucleotide primers specific for T. brucei or T. vivax. The PCR products unique to either T. brucei or T. vivax were identified following amplification of DNA from the blood samples of infected cattle, whereas none was detectable in the DNA from the blood of the cattle exposed to non-infected teneral tsetse. In a concurrent set of experiments, one of the oligonucleotide primers in each pair was biotinylated for use in PCR-ELISA to examine all the blood samples with this assay. Both the PCR and the PCR-ELISA revealed trypanosome DNA in 85% of blood samples serially collected from the cattle experimentally infected with T. brucei. In contrast, the parasitological assays showed trypanosomes in only 21% of the samples. In the blood samples from cattle experimentally infected with T. vivax, PCR and PCR-ELISA revealed trypanosome DNA in 93 and 94%, respectively. Microscopy revealed parasites in only 63% of the BCs prepared from these cattle. Neither PCR nor PCR-ELISA detected any trypanosome DNA in blood samples collected from the animals in the trypanosome-free areas. However, both assays revealed the presence of trypanosome DNA in a number of blood samples from cattle in trypanosomosis-endemic areas.