Patient and professional engagement in the procurement of dental services.

OBJECTIVE: To describe the process of patient and professional engagement in the procurement of a specialist dental service. Stakeholder engagement is the practice of interacting with, and influencing, project stakeholders to the overall benefit of the project and its advocates. The successful completion of a project usually depends on how the stakeholders view it. Their requirements, expectations, perceptions, personal agendas and concerns will influence the project, shape what success looks like, and impact the outcomes that can be achieved. An engagement working group developed a work plan and communications strategy. Following initial scoping the engagement method and resources were developed. Engagement included patient and dentist questionnaires, interviews with specialists and hospital consultants, briefing papers and market engagement workshops. OUTCOME: Feedback from stakeholders provided reassurance of good access and quality, with 98% of patients expressing satisfaction with their overall treatment. Challenges included communication and administrative support. Learning points were diffusion of tension, developing a relationship based on trust, ensuring that stakeholder views are seen to be included in decision making and that there is flexibility in the engagement process. CONCLUSIONS: Good stakeholder engagement with an effective communications strategy is important in the procurement of dental services. Stakeholder engagement should aim to inform, involve, consult, collaborate and empower.

Dose-response effects of diclazuril against pathogenic species of ovine coccidia and the development of protective immunity.

Twin lambs at pasture with their ewes, were divided into seven groups of 10 lambs. One group of 10 lambs served as a non-infected, untreated control. Five groups of 10 lambs were infected with 10,000 oocysts of Eimeria crandallis and 10,000 oocysts of Eimeria ovinoidalis when they were 3 weeks old (day 21 of the study). This produced a good level of infection with high oocysts production and diarrhoea in the lambs. Fourteen days after the primary, artificial challenge (day 35) four of these groups were treated with oral diclazuril at 0.25, 1.0, 2.0 or 4.0mg/kg. Diclazuril treatment was highly effective, dramatically reducing symptoms of diarrhoea and reducing faecal oocyst output by 79.7%, 97.3%, 99.4% and 99.5% respectively in the treated groups within four days. Two weeks post-treatment, and 28 days after the primary coccidial challenge (day 49 of the study), five groups of lambs were re-challenged with 100,000 oocysts of E. crandallis and 100,000 oocysts of E. ovinoidalis (secondary challenge). A group of lambs which had received neither the primary coccidia infection, nor drug treatment (susceptible controls) were also given the secondary challenge. All lambs given the secondary challenge produced high numbers of coccidia and exhibited varying degrees of diarrhoeic faeces. The lambs, which had previously received the higher doses of diclazuril at 2.0 and 4.0mg/kg, developed clinical signs of coccidiosis. These lambs were completely susceptible despite having received the early primary immunising infection of coccidia on day 21. The effects of the secondary challenge were more severe in the groups dosed with the two highest levels of diclazuril than in the susceptible control lambs, which had presumably been exposed to continued low levels of pasture contamination and had acquired a limited degree of immunity from this exposure. It would appear that treatment at the higher dose levels not only eliminated most of the oocysts from the primary challenge but also adventitious infection derived from the grazing paddocks. In contrast, lambs which had received the two lower drug levels of diclazuril (0.25 and 1.0mg/kg) whilst producing large numbers of oocysts, had only transient diarrhoea following secondary challenge. It was concluded that when used as a metaphylactic treatment, diclazuril works rapidly and is effective within four days of administration. Overall, a single dose of diclazuril at either 0.25-1.0mg/kg appears to be highly effective in the control of coccidiosis in young lambs at pasture whilst allowing the development of protective immunity against subsequent heavy coccidia challenge.

Histopathological observations on the activity of diclazuril (Vecoxan) against the endogenous stages of Eimeria crandallis in sheep.

Doses of sporulated oocysts of Eimeria crandallis were administered to 60 housed lambs aged 3-4 weeks that had been raised coccidia-free. Thirty of the lambs were medicated with diclazuril at intervals over a 20-day period post-infection with the remaining lambs serving as untreated controls. Lambs were euthanased between 5 and 22 days post-infection (dpi) and sections of the small intestine and caecum examined histologically. Untreated lambs showed loss of surface epithelial cells and villous atrophy associated with first-generation meronts, crypt destruction and crypt hyperplasia associated with pro-gamont stages. Diclazuril appeared to have a direct effect on several stages of the parasite life cycle, in particular, the large first-generation meront. Indications were that the drug also had an effect on second-generation meronts and gamont stages. Therapeutic benefits of diclazuril treatment appeared greatest when given early in the infection before damage to the intestine occurs although removal of coccidial stages did appear to reduce the pathology of the disease.

A new method for the experimental production of necrotic enteritis and its use for studies on the relationships between necrotic enteritis, coccidiosis and anticoccidial vaccination of chickens.

A new method for the experimental production of necrotic enteritis in chickens is described. The main features are the use of a diet high in wheat and fish meal content; oral administration of a non-lethal inoculum of the coccidium Eimeria maxima followed 6 days later by the bacterium Clostridium perfringens type A per cloaca, so that the bacterial inoculum is deposited at the time and place when and where the intestinal coccidial lesions are maximal; grading of coccidial and clostridial lesions in individual birds sampled during the 14 days following the coccidial infection. The new method was used to examine the relationship between clostridial and coccidial infections. Frank coccidiosis, caused by virulent E. maxima, exacerbated the lesions of necrotic enteritis and other clinical effects due to a subsequent challenge with virulent C. perfringens type A. Immunization with a live, pentavalent, attenuated anticoccidial vaccine (Paracox-5) protected against a severe challenge with heterologous E. maxima. Furthermore, vaccination with Paracox-5, by virtue of its protection against clinical coccidiosis due to the E. maxima challenge, indirectly protected birds against a subsequent challenge with virulent C. perfringens. The results are reconciled with previous field observations on concomitant coccidiosis and necrotic enteritis in chicken flocks.

A novel method for counting eimerian oocysts at very low concentrations in aqueous suspensions.

A novel method for counting eimerian oocysts in samples of drinking water has been developed to fulfil the need for monitoring the delivery of very low concentrations of live anticoccidial vaccines to poultry via pipeline nipple-drinker systems. Advantages of the method are the ease of sample collection and processing, high degrees of accuracy and precision, and a sensitivity of one oocyst ml(-1). Results of a validation test are presented, with a protocol for the method and notes on its use. The coefficient of variation (CoV) of 10 sets of oocyst counts with nominal means of 10 to 160 oocysts ml(-1)ranged from about 16 per cent down to 6 per cent. The recovery efficiency for all 100 validation counts averaged 100.2 per cent with a range of 70-130 per cent. A practical example of field use of the method is given, including a modification to decrease the time taken for counting. In this case, when oocysts were pumped around a pipeline circuit of 129 m for 2.5 hours, the CoV of a mean of 112 oocysts ml(-1)(n = 10) was 12.4 per cent.

The nitroreductase/CB1954 combination in Epstein-Barr virus-positive B-cell lines: induction of bystander killing in vitro and in vivo.

Epstein-Barr virus (EBV)-based gene delivery vectors that preferentially express toxic genes in EBV-infected cells could be used to target EBV-positive tumors for destruction. We have shown previously that the cytosine deaminase (CD) enzyme, which converts the prodrug 5-fluorocytosine (5-FC) into the toxic compound 5-fluorouracil efficiently kills EBV-positive cells in the presence of 5-FC, with a substantial bystander killing effect in vitro and in vivo. To identify the optimal enzyme/prodrug combination for treating EBV-positive lymphomas, we have compared the effectiveness of the CD/5-FC combination with the nitroreductase (NTR)/CB1954 combination for killing EBV-positive B-cell lines. NTR metabolizes CB1954 into an alkylating agent that cross-links DNA. When the CD gene or the NTR gene were transfected into two different EBV-positive B-cell lines in vitro, approximately 90% of cells were killed in a prodrug-dependent manner, although the transfection efficiency was <5%. However, severe combined immunodeficient mouse tumors containing either 30% or 100% of NTR-expressing Burkitt lymphoma (Jijoye) cells were growth inhibited, but not cured, by treatment with intraperitoneal CB1954 (20 mg/kg/day) for 10 days. These results suggest that the NTR/CB1954 combination induces efficient bystander killing of EBV-positive B-cell lines in vitro but may not be as effective as the CD/5-FC combination for treating B-cell lymphomas in vivo.

A new protocol for a challenge test to assess the efficacy of live anticoccidial vaccines for chickens.

The search for vaccines to control coccidioses caused by Eimeria species in chickens (Gallus gallus) is intensifying because of the increasing threat of drug resistance to anticoccidial agents. It is important, therefore, to develop a reliable standard method for the assessment of multivalent vaccine efficacy, because many criteria generally used to judge drug efficacy are not appropriate for vaccines. The lack of correlations between oocyst production, severity of lesions and bird weight gains is discussed. Furthermore, not all Eimeria species cause pathognomonic lesions. A new protocol for a vaccine efficacy test is described which uses growth rate of chickens after virulent challenge as the primary criterion and feed conversion ratio as the secondary criterion for protection against each of the separate coccidioses caused by the seven species of Eimeria that parasitize the chicken. The benefits to this protocol over previous ad hoc experimental designs are: (1) immunization is carried out with multivalent vaccines of Eimeria species up to the maximum of seven that may infect chickens; (2) assessments of immunity are carried out for each species separately so results can not be confounded; (3) the criteria of efficacy are those that are crucial to demonstrate commercial usefulness; (4) the possibility of drawing erroneous conclusions based upon inappropriate criteria such as oocyst production or lesion scores is avoided; (5) because the same criteria are used for each species, direct comparisons may be made amongst immunities to all of the species in the vaccine being tested. Results are presented from tests of three commercial batches of Paracox attenuated anticoccidial vaccine, showing that separate virulent challenges with all seven Eimeria species were controlled in vaccinated chicks.

The pathogenesis of experimental infections of Cryptosporidium muris (strain RN 66) in outbred nude mice.

Three groups of six-week-old nude outbred mice were orally infected with 400, 20,000 and 1,000,000 oocysts of Cryptosporidium muris (strain RN 66) per mouse, respectively. Oocysts were detected in the faeces from 10-18 days post-infection (p.i.) and continued to be shed in large numbers in all groups until the termination of the trial on day 89 p.i. Clinical signs were not observed in any of the infected mice and there was no significant effect on weight gain compared to uninfected controls. Histological examination revealed the presence of parasites confined to the glandular stomach. Parasitised gastric glands were dilated, hypertrophied and filled with numerous parasites. The glands had lost their normal cellular architecture and were lined with many undifferentiated cells. In some mice receiving the largest innoculum, the glandular mucosa was congested and the lamina propria infiltrated with eosinophils, polymorphs and lymphocytes.

Review article: coccidiosis of domestic ruminants.

Coccidiosis in ruminants results from complex interactions between hosts and parasites with many external factors influencing the severity of disease. The study of ruminant coccidiosis is still very much in its infancy despite the advances made in recent years in determining life cycles, pathogenesis, epidemiology and control. Many of the life cycles of the species, considered to be of lesser importance, have still to be elucidated and controversy still exists concerning the pathogenicity of some of these species. Outbreaks of coccidiosis occur with ever increasing frequency yet few drugs exist for prevention. Only limited investigations have been undertaken on immunological methods of control. This paper seeks to review the current knowledge on the biology and control of the coccidial species affecting cattle, sheep and goats.

Variations in oocyst output associated with Cryptosporidium baileyi infections in chickens.

Groups of birds, aged 2, 16 and 30 days, were experimentally infected with 400, 20,000 or 1,000,000 oocysts of Cryptosporidium baileyi. The faecal oocyst output for each group was determined by collection of faeces on Days 0, 5, 7 and 13-21 post-infection (p.i.). The duration of patency of infection, as determined by oocyst excretion, declined more rapidly in the older age groups but persisted longer in birds challenged with the lower oocyst numbers.

Detection of Cryptosporidium parvum using a specific polymerase chain reaction.

The design and use of polymerase chain reaction primers and probes as reagents for the detection of Cryptosporidium parvum are described. Sensitive and specific amplification of a 329 base pair product was demonstrated by ethidium bromide staining and hybridisation of radiolabelled probes. These reagents have the potential for application to diagnostic samples, environmental monitoring and epidemiological surveys.

Giardiasis in lambs at pasture.

Faecal samples from a group of lambs at pasture were screened at weekly intervals for nine weeks for the presence of Giardia species using a modified zinc sulphate flotation method. Fifty-nine of 86 lambs (68.6 per cent) excreted giardia cysts on one or more occasions. They were first detected at approximately three weeks of age and the highest incidence of excretion of cysts occurred when the lambs reached a mean age of 37 days. The lambs had diarrhoea but it was attributed to the presence of Eimeria ovinoidalis.

Immunisation of lambs against coccidiosis.

Two groups of twin lambs, kept with their dams at pasture, were given 10,000 oocysts of Eimeria crandallis and 10,000 oocysts of E ovinoidalis either at birth only, or on four occasions at weekly intervals. A further group received 1000 oocysts of each species three times a week in a 'trickle infection' from birth to 21 days of age. All these lambs, together with a susceptible control group were challenged with 100,000 oocysts of each species at 28 days of age. A fifth group received no inoculations throughout. Bodyweight, faecal consistency and oocyst output were monitored up to nine weeks of age. There was no clinical response to any of the immunising inoculations and no change in the faecal consistency, but the group infected at birth grew significantly faster than the uninfected controls. The pattern of oocyst output showed that only E crandallis developed fully in the newborn animal, but both species multiplied in seven-day-old lambs. The challenge infection produced 80 per cent mortality in the susceptible control group and 20 per cent mortality in the group which had received only one immunising dose at birth. The other immunised groups were well protected and gained more weight than the unchallenged controls. At nine weeks of age, the weight gain of the lambs which had received the 'trickle infection' was significantly higher than that of all the other groups.

Analysis of oocyst wall and sporozoite antigens from three Cryptosporidium species.

A comparison was made of the antigenic composition of oocyst walls and sporozoites from Cryptosporidium baileyi from turkeys, C. muris from rodents, and C. parvum from ruminants, employing immunoblotting and immunofluorescence. In immunoblotting, oocyst antigens were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting (immunoblotting) and detected with rabbit polyclonal anti-C. muris or -C. parvum antibodies or murine monoclonal antibodies developed against C. parvum. Immunofluorescence was used to investigate the reactivity of these monoclonal antibodies with air-dried excystation mixtures of sporozoites and oocysts of the different species. The results from both types of experiment indicated that the three Cryptosporidium species could be differentiated immunologically. In comparison, few antigenic differences were found between a number of isolates of C. parvum in immunoblotting. There was also evidence to suggest that C. parvum and C. baileyi were more closely related antigenically to one another than to C. muris.

Comparison of resistance level and circulating IgG response in chickens experimentally inoculated with a multiple or single immunizing doses of Eimeria acervulina.

Two immunizing methods (Trickle or single immunizing doses) against E. acervulina were tested in chickens. The effects of immunization and challenge upon growth, oocyst output and circulating antibodies response (IgG) were compared. Neither immunization method produced pathogenic effects, similar numbers of oocysts were produced, and the levels of IgG in serum were similar and low in each case. After the challenge, immunized birds showed a high level of resistance but susceptible controls produced very large number of oocysts and showed a marked reduction in the growth. Birds immunized by a trickle infection produced oocysts on two days only and the total number of oocysts per bird was very low, whereas those immunized by a single infection produced oocysts over a period of nine days and the total number of oocysts was higher. Susceptible and birds immunized by a single inoculation showed similar IgG concentration and these were statistically higher than birds immunized by a trickle infection. In susceptible birds the kinetic of IgG was delayed about 4 days.

Observations on the endogenous stages of Eimeria crandallis in domestic lambs (Ovis aries).

Lambs reared coccidia-free were inoculated orally with various numbers of sporulated oocysts of E. crandallis and were killed between 1 and 22 days after inoculation; tissues were examined histologically. Sporozoites were seen 1, 2 and 3 days after inoculation (DAI) in crypt epithelial cells in the mid-jejunum. Infected cells migrated into the lamina propria where the parasite within them developed into a first-generation meront containing about 250,000 merozoites at 10 DAI. A second generation of meronts was seen at 10-12 DAI, each containing up to about 10 merozoites, situated mainly at the bases of crypts in the jejunum and ileum but also in the caecum. From 11 DAI pro-gamonts were seen which were enveloped by the host cell nucleus and which divided in synchrony with the host cell for an undetermined number of generations. Mature gamonts began to develop from them by 16 DAI. Oocyst output began at 16 DAI and rose to a peak at about 22 DAI. Up to 10(8) oocysts were produced per oocyst inoculated. They showed wide variation in size and colour.

Interaction between coccidia and Nematodirus battus in lambs on pasture.

Five thousand oocysts of each of two species of coccidia, Eimeria crandallis and E ovinoidalis or 30,000 infective larvae of Nematodirus battus, given as single infections to three- to five-week-old lambs, caused only transient diarrhoea and had no effect on growth. Lambs infected first with coccidia and two weeks later with N battus suffered severe diarrhoea, weight loss and some deaths. Simultaneous administration of the coccidia and the nematodes increased the clinical severity of the syndrome and increased the numbers of nematode eggs produced.

Ovine coccidiosis: studies on the pathogenicity of Eimeria ovinoidalis and E. crandallis in conventionally-reared lambs, including possible effects of passive immunity.

No pathogenic effect was detected in lambs when 10(4) oocyts of each species were inoculated before 72 h of age. At 4 weeks of age the combined inoculum caused diarrhoea and weight loss, the severity being roughly proportional to the size of the inoculum. Even 1000 oocysts of each species caused diarrhoea; the pathogenic effect was attributable mainly to E. ovinoidalis. Hyperimmunization of ewes during pregnancy (by repeated inoculation of massive doses of oocysts) reduced the effects of oocyst inoculation in their progeny. Levamisole administration during pregnancy increased the birthweight of lambs.