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BACKGROUND: Demand for viral load (VL) monitoring is expected to increase; however, implementation of the multifaceted VL testing poses numerous challenges. We report experiences from Médecins Sans Frontiéres (MSF) and partners in the scale-up of HIV VL in collaboration with the Ministry of Health and Child Care (MoHCC) of Zimbabwe. METHODS: A retrospective data review of routine reports from MSF-supported health facilities in Manicaland Province (Zimbabwe) was conducted. These secondary aggregate data were triangulated, and emerging themes of lessons learnt from VL monitoring were shared. RESULTS: A VL testing coverage of 63% (5966/9456) was achieved among the 40 health facilities, together with a switch rate to second-line antiretroviral therapy (ART) of 46.4% (108/233). The key enablers to scaling-up the VL monitoring were well-equipped and supported VL laboratories, the operationalisation of the on-the-job clinical mentoring and systematic weaning off of better performing health facilities. Concerted efforts from different implementing partners and funders in the HIV programme, and close collaboration with MoHCC were pivotal. CONCLUSION: Our experience indicates that clinical mentoring is effective, and resulted in high VL testing coverage and up-skilling primary health care workers in VL monitoring. Attention must be focused on innovations for improving VL result utilisation, especially the identification and management of patients who fail ART.

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There has been an increase in the introduction of game animals, including African buffaloes, into the Highveld of Zimbabwe to establish private game reserves on condition that they are confined in separate and secured paddocks. Owing to shortages of pastures cattle were grazed in buffalo-grazed paddocks resulting in outbreaks of buffalo-derived theileriosis. This paper reports the results of epidemiological observations carried out on two game reserves to assess the risk of buffalo translocation. The infection rate with Theileria parasites in ticks collected from buffalo-grazed pastures was high and produced fatal theileriosis in susceptible cattle. Similarly, adult R. appendiculatus ticks artificially fed as nymphs on the buffaloes produced fatal infections in susceptible cattle. Theileria parva (Boleni), the vaccine used to immunize cattle against theileriosis, and a buffalo-derived T. parva stabilate (BV-1) were inoculated in naïve buffaloes to study the Theileria carrier-state in these animals. The two buffaloes that had received the Boleni stabilate showed no clinical theileriosis reaction; however, the ticks derived from them produced a subclinical reaction in one susceptible calf. The buffalo which had received stabilate BV-1 developed fever, high schizont parasitosis for 10 days and 15% piroplasms parasitemia. R. appendiculatus ticks fed as nymphs on this buffalo produced fatal theileriosis reaction in a susceptible calf.

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Anaplasma marginale is the causative agent of bovine anaplasmosis, a disease which can be protected by vaccination with the less pathogenic Anaplasma species, A. centrale. Currently, there is no polymerase chain reaction (PCR) assay available which differentiates between different species of Anaplasma or which can differentiate isolates of A. marginale within outbreaks and between different countries. A molecular test specific for A. marginale would be ideal for the identification of Anaplasma species in wild ruminants, as possible reservoirs of anaplasmosis, and to differentiate between A. marginale from A. centrale. A PCR assay was designed to amplify the major surface protein 1alpha gene of the rickettsial bovine pathogen, A. marginale both as an inter- and intra-specific test. The test did not amplify A. centrale or A. ovis, and discriminated A. marginale by amplifying repeat regions within the msp1alpha gene which vary in number between many isolates. The nested A. marginale amplicons varied in size from 630 to 1190bp representing one to eight internal repeats. All 22 Australian isolates tested amplified a 630bp product (one repeat) in contrast to all 19 non-Australian isolates tested. Eight sequences from Australian isolates from different geographical regions confirmed the conserved nature of the Australian A. marginale msp1alpha genes. The Australian 'repeat unit' MSP1a deduced amino acid sequence has been designated as Australian type 1. The msp1alpha PCR method developed here enabled the amplification and comparison of A. marginale isolates originating from North and South America, Africa, Israel and Australia. The method is sensitive and specific for A. marginale. Although additional msp1alpha products were amplified from at least two Australian isolates, the results suggest limited introduction of A. marginale into Australia.

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Fifty-nine Hereford cattle susceptible to tick-borne diseases were used as tracer animals to assess the tick challenge and pathogenicity of Theileria parva under field conditions in Zimbabwe. They were moved periodically in groups of five to three commercial farms (one group consisted of four) during seasons of Rhipicephalus appendiculatus nymphal and adult activity. All tracer cattle were herded together with the farm cattle but were not dipped. The nymphal tick counts were high on two of the farms (up to 2000 per animal) but were very low on the third farm (less than ten per animal). On the three farms, 19 out of 24 (76%) tracers had patent Theileria schizonts. There was a range of clinical manifestations of theileriosis with acute and fatal infections occurring on one farm. The adult R. appendiculatus infestations during the wet season numbered 120-800 per animal on the three farms. The disease transmitted by the adults was very pathogenic on the three farms; 30 out of 35 (86%) had severe theileriosis infections. Cattle, which survived the nymphal diseases challenge, showed various degrees of immunity to subsequent T. parva challenge transmitted by adult ticks. Therefore, 13 out of 18 (72%) of these cattle had a second disease episode and the case fatality rate on the three farms was 46%. The factors which determined the epidemiological status of Theileria challenge on the farms, such as the farming systems and presence of wild animals, are discussed.

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Eight cattle immunized with cattle-derived Theileria parva Boleni stabilate together with six susceptible controls were released in Dombawera Game Park on the Highveld of Zimbabwe. This coincided with Rhipicephalus appendiculatus nymphal activity. The cattle grazed together with African buffaloes (Syncerus caffer) and were not treated against tick infestation. The nymphal tick infestation was high, and seven of the eight immunized cattle and three of the controls had severe and fatal reactions. Subsequently, two stocks of Theileria parva to be tested for their immunizing abilities were prepared-one from adult ticks which were fed as nymphs on one of the sick control animals (Dom 268) and the other from adult ticks collected from pastures grazed by buffaloes (Bv-1). Two groups of cattle were immunized with either the Dom 268-derived strain (eight animals) or the Bv-1-derived strain (four animals). These together with three non-immunized controls, were released in Bally Vaughaun Game Park in the Highveld, where buffaloes are present, during the season of nymphal tick activity. A third group of five cattle, immunized with stabilate Bv-1, and three non-immunized controls were released at the same site during the season of adult tick activity. The nymphal and adult tick infestations of the cattle were large and more than 2000 nymphs and 1000 adult ticks were counted per animal. Cattle were treated with a pyrethroid pour-on preparation to control the tick infestation and screw-worm strike. The immunized cattle in the three groups survived the theileriosis challenge for a period of 18 months, but the non-immunized control cattle suffered a severe and fatal theileriosis 19-23 days after being placed on the pasture.

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The Theileria parva carrier-state in cattle on commercial farms on Zimbabwe was investigated using parasitological and serological methods. The proportion of cattle showing Theileria piroplasms on two farms, which had recent histories of disease outbreaks, were 64% (n = 106, total of heifers and weaned calves examined) and 71.5% (n = 60) while the proportion of T. parva antibodies for the same animals were 59% and 98.5%, respectively. On four farms where no cases of the disease occurred for over 10 years, the average proportion of animals showing piroplasms and antibodies were 55.4% (range 32-82, n = 223) and 73% (range 47-91, n = 223), respectively. However, on another three farms which had no history of theileriosis outbreaks these proportions were very low, being 11.4% (0-24, n = 157) for piroplasms and 12.2% (5-23, n = 157) for antibodies. The mean infection rate in unfed Rhipicephalus appendiculatus adults collected from farms with a high prevalence of cattle which were carriers of Theileria piroplasms during the tick activity season was 29% (range 12-60%) with 9.3 (range 2-18.7) mean infected acini per infected tick. The infectivity of different tick batches to susceptible cattle produced a wide spectrum of theileriosis reactions. Laboratory controlled experiments were carried out to study the persistence of T. parva (Boleni) piroplasms in cattle immunized with this strain as well as its infectivity for ticks and its subsequent transmissibility to cattle. Examination of the salivary glands of 15 batches of ticks collected from six immunized cattle on three different occasions over 18 months showed that none were infected with Theileria parasites. However, the infectivity of other ticks in the same batches to susceptible animals was demonstrated 6, 10 and 18 months after cattle had been immunized with Boleni stabilate.

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Transmission of a Babesia species to susceptible cattle by Rhipicephalus evertsi evertsi ticks picked off a sable which died of acute babesiosis is described. The parasite was initially isolated by feeding R. e. evertsi ticks on a susceptible, splenectomised bovine which developed parasitaemia. Blood stabilate from the parasitaemic bovine produced a fatal babesiosis in a spleen--intact bovine. Clinical signs shown by the affected animals corresponded with those of acute babesiosis. Parasitological examination, the immunofluorescence antibody test and the polymerase chain reaction test revealed that the parasite transmitted by the ticks initially and the blood stabilates prepared from affected animals was Babesia bigemina. This parasite was morphologically identical to that observed in Giemsa-stained blood smears prepared from the dead sable.

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Theileria parva (Boleni) stock from Zimbabwe was used to immunise 24 susceptible Friesian calves by infection and treatment with oxytetracycline. Twenty-eight days after immunisation the animals in groups of 6 plus 2 susceptible controls were subjected to challenge: 3 groups with heterologous stocks and one group with the immunising stock. Theileria parva (Boleni) gave good protection against homologous challenge, the T. parva (Muguga, Kenya) and buffalo-derived T. parva (Serengeti transformed, Tanzania) parasite stocks. It did not protect against the T. parva (Kasoba, Malawi) stock and 3 out of 6 immunised cattle died and the remaining 3 had to be treated with parvaquone. In a second experiment, the 6 T. parva (Boleni) immunised animals which had received homologous challenge, together with the 2 controls which had recovered without treatment from T. parva (Boleni) infection, were challenged with the T. parva (Kasoba) stock. Four out of 6 of the immunised animals resisted the challenge with mild to moderate reactions. The other 2 animals had severe reactions and one died. The 2 control animals which recovered from T. parva (Boleni) infection resisted the T. parva (Kasoba) challenge and both had mild reactions. It is suggested that oxytetracycline used in the first experiment may have interfered with the expression of the full protective capacity against the virulent T. parva (Kasoba) stock. Further studies on the use of the T. parva (Boleni) stock without oxytetracycline treatment could identify a more broadly immunising effect and a more economical vaccination method.

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Forty Theileria schizont-infected lymphocyte culture isolates from Zimbabwe were characterized using a panel of antischizont monoclonal antibodies (MAbs) and 4 Theileria parva DNA probes containing cloned extrachromosomal element, Tpr repetitive, ribosomal and telomeric sequences. The Theileria isolates were assigned as T. parva or T. taurotragi on the basis of reactivities with MAbs and restriction fragment length polymorphisms (RFLPs) detected using the extra-chromosomal element probe. Cattle-derived T. parva isolates were relatively homogeneous on the basis of reactivities with MAbs and RFLPs detected using Tpr repetitive and ribosomal DNA probes. In contrast to previous results from Kenya, most of the cattle-derived isolates from Zimbabwe exhibited very similar Tpr restriction fragment patterns, although the Tpr genotypes of buffalo-derived isolates were heterogeneous. This suggests that selection for a particular Tpr genotype may be occurring in cattle. Many isolates with similar Tpr genotypes were differentiated by RFLPs detected using the telomeric DNA probe. The T. parva Boleni immunizing stock was distinguished from all other isolates by telomeric RFLPs. The T. parva Boleni Tpr repetitive DNA probe cross-hybridized with T. taurotragi DNA and detected RFLPs between different T. taurotragi isolates.

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The seroconversion by indirect ELISA to Cowdria ruminantium over the first year of life of sixty-six Malawi zebu calves born into groups which were dipped 17 times per year was compared to seroconversion of 32 calves born into non-dipped groups. Amblyomma variegatum tick counts and clinical disease in each group of cattle were monitored throughout the study period. No cases of heartwater were seen in either group of calves over the first 22 months of life. Only one case of heartwater was observed, in an 8 year old cow, in the 1,800 intensively monitored cattle over the same period. By 12 months of age almost all undipped calves had seroconverted and 50% of seroconversions were attributed to nymphal challenge. In contrast, only 41% of calves had become seropositive by 12 months of age in the dipped groups. The dipping regime used therefore significantly decreased seroconversion rates to C. ruminantium in these calves. 73% of calves had detectable levels of maternal antibodies to C. ruminantium in the first 4 weeks of life. Antibody levels in each of the calves in dipped groups had waned to below the cut off point for the ELISA by 8-12 weeks. Seroconversion did not occur in the first 8-12 weeks of life in dipped herds. The indirect ELISA test results were not significantly different in the proportion positive in single tests at 12 months of age, or by cumulative test results of the previous 9 months, and therefore the test may be of value as a test of herd immunity.(ABSTRACT TRUNCATED AT 250 WORDS)

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Data collected in the Zimbabwean province of Mashonaland-West, in the period 1980-1988, showed that mortality in calves owing to Theileria parva bovis infection (January disease) was significantly lower in animals younger than 7 months than in older cattle. Groups of seven Holstein-Friesian calves from non-immune dams aged approximately 1, 4, 7, 10 and 13 months were infected with a Theileria parva bovis tick-derived stabilate. The dose chosen was lethal for 40% of the calves in the trial. Mortality was highest in the 4-month age group. The reactions in the 7-, 10- and 13-month age groups became progressively milder. The reactions in the 1-month old calves were the least marked, being very mild. The age-related resistance in the youngest calves, as can be concluded from our results, is only of short duration and cannot explain the lower incidence of January disease observed in calves in the field.

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