What is the real number of Lyme disease cases in Canada?

BACKGROUND: Lyme disease is emerging in Canada due to expansion of the range of the tick vector Ixodes scapularis from the United States. National surveillance for human Lyme disease cases began in Canada in 2009. Reported numbers of cases increased from 144 cases in 2009 to 2025 in 2017. It has been claimed that few (< 10%) Lyme disease cases are reported associated with i) supposed under-diagnosis resulting from perceived inadequacies of serological testing for Lyme disease, ii) expectation that incidence in Canadian provinces and neighbouring US states should be similar, and iii) analysis of serological responses of dogs to the agent of Lyme disease, Borrelia burgdorferi. We argue that performance of serological testing for Lyme disease is well studied, and variations in test performance at different disease stages are accounted for in clinical diagnosis of Lyme disease, and in surveillance case definitions. Extensive surveillance for tick vectors has taken place in Canada providing a clear picture of the emergence of risk in the Canadian environment. This surveillance shows that the geographic scope of I. scapularis populations and Lyme disease risk is limited but increasing in Canada. The reported incidence of Lyme disease in Canada is consistent with this pattern of environmental risk, and the differences in Lyme disease incidence between US states and neighbouring Canadian provinces are consistent with geographic differences in environmental risk. Data on serological responses in dogs from Canada and the US are consistent with known differences in environmental risk, and in numbers of reported Lyme disease cases, between the US and Canada. CONCLUSION: The high level of consistency in data from human case and tick surveillance, and data on serological responses in dogs, suggests that a high degree of under-reporting in Canada is unlikely. We speculate that approximately one third of cases are reported in regions of emergence of Lyme disease, although prospective studies are needed to fully quantify under-reporting. In the meantime, surveillance continues to identify and track the ongoing emergence of Lyme disease, and the risk to the public, in Canada.

A comparison of exposure to risk factors for giardiasis in non-travellers, domestic travellers and international travellers in a Canadian community, 2006-2012.

The purpose of this study is to determine how demographic and exposure factors related to giardiasis vary between travel and endemic cases. Exposure and demographic data were gathered by public health inspectors from giardiasis cases reported from the Region of Waterloo from 2006 to 2012. Logistic regression models were fit to assess differences in exposure to risk factors for giardiasis between international travel-related cases and Canadian acquired cases while controlling for age and sex. Multinomial regression models were also fit to assess the differences in risk profiles between international and domestic travel-related cases and endemic cases. Travel-related cases (both international and domestic) were more likely to go camping or kayaking, and consume untreated water compared to endemic cases. Domestic travel-related cases were more likely to visit a petting zoo or farm compared to endemic cases, and were more likely to swim in freshwater compared to endemic cases and international travel-related cases. International travellers were more likely to swim in an ocean compared to both domestic travel-related and endemic cases. These findings demonstrate that travel-related and endemic cases have different risk exposure profiles which should be considered for appropriately targeting health promotion campaigns.

Comparison of calculation methods used for the determination of anthelmintic resistance in sheep in a temperate continental climate.

This study compared results obtained with five different fecal egg count reduction (FECR) calculation methods for defining resistance to ivermectin, fenbendazole, and levamisole in gastrointestinal nematodes of sheep in a temperate continental climate: FECR1 and FECR2 used pre-and posttreatment fecal egg count (FEC) means from both treated and control animals, but FECR1 used arithmetic means, whereas FECR2 used geometric means; FECR3 used arithmetic means for pre- and posttreatment FECs from treated animals only; FECR4 was calculated using only arithmetic means for posttreatment FECs from treated and control animals; and FECR5 was calculated using mean FEC estimates from a general linear mixed model. The classification of farm anthelmintic resistance (AR) status varied, depending on which FECR calculation method was used and whether a bias correction term (BCT, i.e., half the minimum detection limit) was added to the zeroes or not. Overall, agreement between all methods was higher when a BCT was used, particularly when levels of resistance were low. FECR4 showed the highest agreement with all the other FECR methods. We therefore recommend that small ruminant clinicians use the FECR4 formula with a BCT for AR determination, as this would reduce the cost of the FECRT, while still minimizing bias and allowing for comparisons between different farms. For researchers, we recommend the use of FECR1 or FECR2, as the inclusion of both pre- and posttreatment FECs and use of randomly allocated animals in treatment and control groups makes these methods mathematically more likely to estimate the true anthelmintic efficacy.

Comparison of calculation methods used for the determination of anthelmintic resistance in sheep in a temperate continental climate.

This study compared results obtained with five different fecal egg count reduction (FECR) calculation methods for defining resistance to ivermectin, fenbendazole, and levamisole in gastrointestinal nematodes of sheep in a temperate continental climate: FECR1 and FECR2 used pre- and posttreatment fecal egg count (FEC) means from both treated and control animals, but FECR1 used arithmetic means, whereas FECR2 used geometric means; FECR3 used arithmetic means for pre- and posttreatment FECs from treated animals only; FECR4 was calculated using only arithmetic means for posttreatment FECs from treated and control animals; and FECR5 was calculated using mean FEC estimates from a general linear mixed model. The classification of farm anthelmintic resistance (AR) status varied, depending on which FECR calculation method was used and whether a bias correction term (BCT, i.e., half the minimum detection limit) was added to the zeroes or not. Overall, agreement between all methods was higher when a BCT was used, particularly when levels of resistance were low. FECR4 showed the highest agreement with all the other FECR methods. We therefore recommend that small ruminant clinicians use the FECR4 formula with a BCT for AR determination, as this would reduce the cost of the FECRT, while still minimizing bias and allowing for comparisons between different farms. For researchers, we recommend the use of FECR1 or FECR2, as the inclusion of both pre- and posttreatment FECs and use of randomly allocated animals in treatment and control groups makes these methods mathematically more likely to estimate the true anthelmintic efficacy.

Factors related to Campylobacter spp. carriage in client-owned dogs visiting veterinary clinics in a region of Ontario, Canada.

From July 2008 until May 2009, 240 client-owned pet dogs from seven veterinary clinics in the Region of Waterloo, Ontario, Canada participated in a study to determine pet-related management factors that may be associated with the presence of Campylobacter spp. in dogs. The prevalence of Campylobacter spp. carriage in our study population of pet dogs was 22%, with 19% of the dogs positive for C. upsaliensis, and 3% positive for C. jejuni. A significant risk factor from multivariable logistic regression models for both Campylobacter spp. and C. upsaliensis carriage was having homemade cooked food as the dog's diet or added to its diet, and a significant sparing factor for both models was treatment with antibiotics in the previous month. Increasing age of the dog decreased the odds of Campylobacter spp. and C. upsaliensis carriage. Based on the high prevalence of Campylobacter, and specifically C. upsaliensis, further research concerning pet dogs as a risk factor for campylobacteriosis in humans is warranted.

Passive immunity in Ontario dairy calves and investigation of its association with calf management practices.

Adequate passive transfer of maternal immunoglobulin is important for optimal health and performance in newborn dairy calves. From June to October 2003 and January to April 2004, blood samples were collected from 961 dairy calves 0 to 8 d of age on 11 farms in southwestern Ontario. This was followed by a second study conducted from May to October 2004, in which similar samples were taken from 422 calves up to 8 d of age on 119 dairy farms throughout southern Ontario. For each sample collected, serum refractometry was used to evaluate serum total protein (TP) as a measure of passive transfer of maternal immunity. During each study, producers were asked to provide information on calf management practices, including details of colostrum feeding. Data were analyzed to assess the levels of maternal immunity present in the calves, and to investigate whether these were associated with any calf management or colostrum feeding practices used on the farms. Serum TP readings ranged from 3.5 to 9.8 g/dL. Controlling for any effects of variation between farms, we found no statistically significant difference in serum TP levels, or risk of failure of passive transfer (FPT), between heifer and bull calves. The odds of FPT in calves on farms where more than 75% of cows were usually allowed to remain with their calves for more than 3 h after calving were significantly higher than the odds of FPT in calves on farms where dams and calves were separated within 3 h of the birth. Furthermore, an increased volume of colostrum given to calves within 6 h of birth was significantly associated with a reduced risk of FPT in calves. Information from this work provides valuable insight into the efficiency of passive transfer in newborn dairy calves in southern Ontario.

Temporal trends in Giardia occurrence in the Grand River and surrounding tributaries, Waterloo, Ontario (2005-2013), a retrospective analysis of surveillance data.

Giardia contamination in the Grand River Watershed (south-western Ontario, Canada) was monitored from 2005 to 2013 as part of FoodNet Canada. Our study objectives were to describe the temporal pattern of Giardia occurrence and determine whether water quality parameters and bacterial indicators could act as effective markers for Giardia occurrence. Water samples were collected monthly from the Grand River near a drinking water intake point (2005-2013) and also collected intermittently from other areas in the watershed during the study period. Samples were tested for Giardia cysts using the US EPA method 1623. Samples were also tested for chemical and microbial water quality indicators. Univariable and multivariable linear regression models were built to examine whether temporal, water quality and bacterial indicators were associated with Giardia cyst concentration. Giardia cysts were identified in 89% of samples (n = 228), with highest measured concentrations downstream of a waste water treatment plant outfall. Year and season were found to be predictors for Giardia occurrence. Concentrations were significantly higher in the winter and fall compared to the summer, and significantly higher in 2007 compared to other study years. After controlling for season, year and sampling location, dissolved oxygen was the only variable significantly associated with Giardia cyst concentration. Seasonal peaks in Giardia cyst concentrations in samples collected near the intake for the drinking water plant did not align with the seasonal peak in human Giardiasis cases in this region that are reported annually by public health authorities. This suggests that the risk of contracting Giardiasis from treated drinking water in this community is possibly low when the treatment plant is functioning adequately. Instead, waterborne exposure is likely the result of seasonal behaviours surrounding recreational water use. Therefore, the collective findings of our study are important to help inform future risk management studies and guide public health protection policies.

Public Health Follow-up of Suspected Exposure to Echinococcus multilocularis in Southwestern Ontario.

In the 3 years since the first report of canine alveolar echinococcosis (AE) in Ontario, three additional cases have been diagnosed in the province. Of the four cases reported to date, three have had no known history of travel outside the province. It is possible that this development is an indication of previously unrecognized environmental contamination with Echinococcus multilocularis eggs in some areas of the province. If so, there is the potential for an emerging threat to human health. This article describes a local public health department's investigation of the possible exposure to E. multilocularis of a number of individuals who had had contact with the latest of the four cases of canine AE, and summarizes a comprehensive decision process that can be used by public health departments to assist in the follow-up of such exposures.

Prevalence of zoonotic agents in dogs visiting hospitalized people in Ontario: implications for infection control.

Visitation of hospitalized people by dogs is becoming commonplace, but little is known about the potential health risks of introducing dogs to healthcare settings. This cross-sectional study evaluated the prevalence of zoonotic agents in a group of 102 visitation dogs from a variety of sources across Ontario. Between May and July 2004, owners were interviewed by a standardized questionnaire while dogs underwent a standardized physical examination. One specimen of faeces, hair-coat brushings and one rectal, aural, nasal, oral and pharyngeal swab were collected from each dog and tested for 18 specific pathogens. All dogs were judged to be in good health. Zoonotic agents were isolated from 80 out of 102 (80%) dogs. The primary pathogen was Clostridium difficile, which was isolated from 58 (58%) faecal specimens. Seventy-one percent (41/58) of these isolates were toxigenic. Extended-spectrum beta-lactamase Escherichia coli was isolated from one (1%) dog, extended-spectrum cephalosporinase E. coli was isolated from three (3%) dogs, and organisms of the genus Salmonella were isolated from three (3%) dogs. Pasteurella multocida or Pasteurella canis was isolated from 29 (29%) oral swabs, and Malassezia pachydermatis was isolated from eight (8%) aural swabs. Giardia antigen was present in the faeces of seven (7%) dogs, while Toxocara canis and Ancylostoma caninum were detected in two (2%) dogs and one (1%) dog, respectively. Methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, Campylobacter spp., Microsporum canis, group A streptococci, Pseudomonas aeruginosa and Cryptosporidium spp. were not detected. Further information is needed before the full implications of these findings for infection control can be assessed properly.

Identification of effective treatment criteria for use in targeted selective treatment programs to control haemonchosis in periparturient ewes in Ontario, Canada.

Haemonchosis is often associated with late gestation and parturition in ewes in Canada. Due to widespread concerns about development of anthelmintic resistance (AR), targeted selective treatment (TST), where individual animals are treated with an anthelmintic rather than the entire flock, is a possible strategy to control clinical signs in recently lambed ewes while still maintaining parasite refugia. Performing fecal egg counts (FEC) on individual animals is often cost-prohibitive, so indicators that identify ewes with high FEC are essential for TST programs. The study objectives were to: a) evaluate the ability of four TST indicators to identify periparturient ewes with high Haemonchus sp. FEC and b) determine appropriate treatment thresholds for statistically-significant indicators. A field study was conducted during the 2013 and 2014 lambing seasons (February-May) on three client-owned farms in Ontario with documented AR and problems with haemonchosis in ewes. Ewes were examined within three days of lambing and selected for treatment with oral closantel (10mg/kg body weight), a novel anthelmintic to Canada, if they met at least one of four criteria: a) the last grazing season was their first grazing season; b) body condition score ≤2; c) Faffa Malan Chart (FAMACHA©) score ≥3; and/or d) three or more nursing lambs. Fecal samples were collected per rectum on the treatment day from each of 20 randomly selected treated and untreated ewes on each farm. Haemonchus sp. percentages on each farm, as determined by coproculture, ranged from 53% to 92% of total fecal trichostrongyle-type egg counts. Mean Haemonchus sp. FECs were significantly higher in treated ewes (n=136) than in untreated ewes (n=103) on the day of treatment in both years (p=0.001), suggesting the indicators were suitable for identifying animals with high Haemonchus sp. FEC. A linear mixed model was fit with logarithmic-transformed Haemonchus sp. FEC as the outcome variable, the four indicators and year as fixed effects, and farm as a random effect. FAMACHA© score was the sole indicator to remain significantly associated with FEC (p=0.002). A receiver-operator curve determined that test sensitivity was maximized (92.4%) with FAMACHA© score ≥3 as the sole indicator. FAMACHA© score should therefore be included in TST programs to identify ewes requiring treatment at lambing due to Haemonchus sp.

Efficacy of closantel against ivermectin- and fenbendazole-resistant Haemonchus sp. in sheep in Ontario, Canada.

In Ontario, Canada, widespread resistance to ivermectin and fenbendazole, the only readily available ovine anthelmintics, has been documented, primarily in Haemonchus sp. In other parts of the world, closantel has been used to control such infections; however, the drug was not currently licensed for use in Canada and the USA. A randomized controlled trial was conducted on six client-owned farms in Ontario in 2013 and 2014 to determine the efficacy of closantel (Flukiver® 5% Oral Suspension, Elanco Animal Health, 10mg/kg bodyweight) against ivermectin- and fenbendazole-resistant Haemonchus sp. infections in periparturient ewes and grazing lambs. Three farms were randomly assigned to treat all ewes, and three farms were randomly assigned to selectively treat individual ewes at lambing, using predetermined criteria. Fecal samples were collected from a minimum of 15 randomly selected ewes and 13 lambs per group on each farm at the time of treatment and approximately 14days later. Trichostrongyle-type fecal egg counts (FEC) were performed using a modified McMaster technique with a lower detection limit of 8.3 eggs per gram of feces (epg). Haemonchus-specific FECs were determined by multiplying FECs by the proportion of Haemonchus sp. identified from coproculture for each farm; Haemonchus-specific FEC reductions were calculated for each farm. Twenty grazing lambs had FECs conducted monthly, and when mean monthly FECs surpassed 200 epg, all lambs were randomly allocated to either closantel, positive control (ivermectin, fenbendazole, or levamisole) or negative control groups. Pre-treatment Haemonchus-specific mean FECs ranged from 27 to 3359 epg in ewes and 0-5698 epg in lambs. Efficacy of closantel against Haemonchus sp. ranged from 99% (95% CI: 97%-99%) to 100% in recently lambed ewes on all farms in both years (total n=274 ewes), and from 99% (95% CI: 98%-99%) to 100% in grazing lambs in both years on all but one farm (total n=171 lambs). On the latter farm, a whole flock treated farm, closantel efficacy in grazing lambs was 84% (95%CI: 81%-88%) in the first year, but 100% in the second year. Levamisole was effective against overall GIN in lambs on only two farms. Ivermectin and fenbendazole resistance continued to be present, particularly in Haemonchus sp. Closantel had excellent efficacy against Haemonchus sp. over the two year study period, regardless of treatment group, and therefore should be considered one viable component of sustainable integrated parasite control programs for farms with documented anthelmintic resistance and problems with haemonchosis.

Risk factors associated with Neospora caninum abortion in Ontario Holstein dairy herds.

The objective of this epidemiological study was to identify risk factors for Neospora caninum-related abortions in Ontario Holstein dairy herds. A total of 88 herds, consisting of 5080 cattle, and utilizing Dairy Herd Improvement (DHI) services, were divided into three groups. Case (n = 30) and first control (n = 31) herds were selected from 1998 and 1999 fetal abortion submissions to the Animal Health Laboratory, University of Guelph, that were histopathologically positive or negative, respectively, for N. caninum. A second control group (n = 27) was selected from multiple sources of herds sampled within the previous 4 years that had a low seroprevalence (<7%) to N. caninum. Between May and December 1999, all available cows on all farms, in parity one or greater, were blood sampled. The sera were then analyzed for antibody to N. caninum using a kinetic ELISA. A survey administered at the time of sampling recorded information on housing, animal species present, manure management, reproduction, biosecurity practices, wildlife observations, peri-parturient cow management, herd disease history and nutrition. Production and other herd parameters were obtained from DHI records. Logistic regression indicated that the following parameters were positively associated with a N. caninum abortion in a herd: the N. caninum herd seroprevalence (OR = 1.1), the total number of dogs on a farm (OR = 2.8), the frequency that dogs were observed defecating in mangers (OR = 2.8), the number of horses on a farm (OR = 3.1), the observed annual rate of retained fetal membranes (OR = 1.2) and the observed annual rate of cows returning to estrus after pregnancy confirmation (OR = 1.2). Factors negatively associated were the frequency of stray cats and wild canids observed on a farm (OR = 0.4 and OR = 0.7, respectively) and the housing of heifers on loafing packs (a housing pen divided into feed manger, scrape alley and bedded pack areas, OR = 0.1).

Pharmacology of diminazene: a review.

Chemotherapy for trypanosomiasis in domestic livestock depends on only a few compounds, of which several are chemically closely related. Of these compounds, the most widely used therapeutic agent in cattle, sheep and goats is diminazene aceturate. Diminazene was first described in 1955. Subsequently, a substantial body of data has been generated on various pharmacological aspects of the compound. In this review, we consider the current status of knowledge concerning the therapeutic spectrum of diminazene, resistance to diminazene in trypanosomes, and combination therapeutic regimens in which diminazene has been administered together with other compounds. Analytical techniques for diminazene, the pharmacokinetics of diminazene, data concerning diminazene's toxicity, and the different molecular mechanisms by which diminazene may exhibit trypanocidal action are also considered.

An in vitro assay for drug sensitivity of Trypanosoma congolense using in vitro-derived metacyclic trypanosomes.

The sensitivity of seven populations of T. congolense to the salts of three trypanocides, diminazene, isometamidium and homidium, were determined in vitro using in vitro-derived metacyclic trypanosomes. The trypanosomes were incubated at 35 degrees C for 48 h with various drug concentrations (0.5 ng-50 micrograms/ml) and then transferred to cultures containing bovine endothelial-cell monolayers, to assess their viability over the following 5 days as compared to control trypanosomes that had been incubated without drug. The sensitivity to each drug was expressed as the minimum effective drug concentration which killed 100% of the trypanosomes in a given population within the 5 days. Using this assay, population IL 1180, characterised as being highly sensitive to all three drugs in vivo, required 10 ng/ml isometamidium chloride, 50 ng/ml homidium bromide or chloride and 5000 ng/ml diminazene aceturate to kill the entire population in vitro. In contrast, two derivatives of IL 1180 in which resistance to isometamidium had been induced in mice, IL 3343 and IL 3344, required isometamidium chloride at a concentration of 1000 ng/ml and 2000 ng/ml, respectively, to eliminate the populations. The in vitro results showed that the increase in level of resistance to isometamidium in these populations was associated with at least a 200-fold increase in resistance in both populations to homidium, but no increase in resistance to diminazene. KE 2887 and CP 81, two isolates expressing high levels of resistance to both isometamidium and homidium in mice and cattle, were both resistant in vitro to isometamidium chloride and homidium salts at 100 ng/ml. Furthermore, while the former population was resistant to 10,000 ng/ml diminazene aceturate, the latter was sensitive to 5000 ng/ml. IL 3274 and IL 3330, characterised as expressing high levels of resistance to all three drugs in vivo, were shown to be resistant to isometamidium chloride and homidium salts at 1000 ng/ml, and to diminazene aceturate at 10,000 ng/ml. Finally, the in vitro IC100 (concentration of drug required to eliminate 100% of the population) results were consistent with the maximum amounts of each drug detected in vivo.

Drug sensitivity screening in vitro of populations of Trypanosoma congolense originating from cattle and tsetse flies at Nguruman, Kenya.

An in vitro assay that utilises in vitro-derived metacyclic trypanosomes was used to determine the drug sensitivity of 7 populations of Trypanosoma congolense collected from cattle and tsetse flies at Nguruman; a trypanosomiasis-endemic area in southwest Kenya. The metacyclic trypanosomes used in the assay were obtained from cultures initiated directly from either the blood of cattle with low levels of parasitaemia or from guts of infected tsetse flies. Sensitivities to isometamidium chloride, diminazene aceturate and homidium salts were assessed at various drug concentrations (0.5 ng-50 micrograms/ml). The results were compared with those obtained with two characterised laboratory populations. In spite of the fact that isometamidium chloride had not been widely used at Nguruman, two of the stocks (KE 3302 and KE 3303) expressed high levels of resistance to this drug (resistant to 100 ng/ml and 1000 ng/ml, respectively). In contrast, all of the populations examined were as sensitive, or more so, to diminazene aceturate than the sensitive laboratory clone IL 1180; two stocks (KE 3305 and KE 3306) were more sensitive (resistant to 0.5 microgram/ml, sensitive to 1 microgram/ml) and three stocks expressed the same level of sensitivity as IL 1180 (resistant to 1 microgram/ml, sensitive to 5 micrograms/ml). Since the results of the in vitro assay correlated well with field observations it was concluded that the assay would be a useful tool in epidemiological studies to determine the resistance phenotypes of trypanosome populations in the field, thereby enabling development of appropriate control measures for particular areas.

Therapeutic and prophylactic activity of isometamidium chloride against a tsetse-transmitted drug-resistant clone of Trypanosoma congolense in Boran cattle.

An investigation was conducted on the therapeutic and prophylactic activity of isometamidium chloride (SamorinR) in Boran (Bos indicus) cattle against a Trypanosoma congolense clone, IL 3270. This clone was derived, without drug selection, from a stock originally isolated in Burkina Faso and has previously been shown to be resistant to isometamidium in both cattle and mice using an infection and treatment regimen. A group of 5 cattle were treated intramuscularly with 1.0 mg kg-1 isometamidium chloride and 28 days later challenged with Glossina morsitans centralis infected with T. congolense IL 3270. All 5 cattle and 17 untreated cattle challenged on the same day became parasitaemic by day 16 post challenge, indicating that prophylaxis did not extend to 28 days post treatment. The cattle were then treated with isometamidium chloride at one of the following doses and by different routes of administration; 1.0 or 2.0 mg kg-1 intramuscularly, 0.25, 0.5, 0.75 or 1.0 mg kg-1 intravenously. Infections relapsed in all cattle at an interval of 12-21 days following treatment, with the exception of those treated with 2.0 mg kg-1 intramuscularly in which the development of relapse infections was delayed. Similar studies were also conducted with a highly sensitive clone of T. congolense, IL 1180. Infections in cattle with this clone were eliminated by intravenous treatment with 0.25 mg kg-1 isometamidium chloride or intramuscular treatment with 0.5 mg kg-1 isometamidium chloride. Thus, although intravenous administration of isometamidium eliminated a fully sensitive infection, treatment by this route appeared not to enhance the therapeutic efficacy of the drug in the treatment of a T. congolense clone which expresses a high level of resistance.

Long-term occurrence of Trypanosoma congolense resistant to diminazene, isometamidium and homidium in cattle at Ghibe, Ethiopia.

Ten trypanosome isolates were collected at random from cattle at Ghibe, Ethiopia, in February 1993 and all shown to be savannah-type Trypanosoma congolense. When inoculated into naïve Boran (Bos indicus) calves, all 10 isolates were resistant to diminazene aceturate (Berenil), isometamidium chloride (Samorin) and homidium chloride (Novidium) at doses of 7.0 mg/kg body weight (b.w.), 0.5 mg/kg b.w. and 1.0 mg/kg b.w., respectively. In order to determine whether this multiple-drug resistance was expressed by individual trypanosomes, clones were derived from two of the isolates and characterised in mice for their sensitivity to the three compounds; by comparison to drug-sensitive populations, the two clones expressed high levels of resistance to all 3 trypanocides. In experiments to characterise the uptake kinetics of [14C]-Samorin, the maximal rates of uptake (Vmax) for 4 Ghibe isolates ranged from 9.2 to 15.0 ng/10(8) trypanosomes/min. In contrast, Vmax for the isometamidium-sensitive clone T. congolense IL 1180 was 86.7 +/- 8.6 ng/10(8) trypanosomes/min. Lastly, molecular karyotypes were determined for eight isolates: seven different chromosome profiles were observed. These data indicate that in February 1993 there was a high prevalence of drug-resistant trypanosome populations with different chromosome profiles in cattle at Ghibe. Since a similar situation existed at the same site in July 1989, this suggests that the drug-resistance phenotype of trypanosomes at Ghibe had not altered over a 4 year period.

Concentrations of isometamidium in the sera of cattle challenged with drug-resistant Trypanosoma congolense.

The relationship between serum concentrations of the prophylactic trypanocidal drug isometamidium chloride and protection against tsetse challenge with two populations of Trypanosoma congolense was investigated in Boran (Bos indicus) cattle, using an isometamidium-ELISA. Isometamidium chloride (Samorin) was administered to cattle at a dose rate of 1.0 mg/kg body weight by deep intramuscular injection. Thereafter, the animals were challenged at monthly intervals with either a drug-sensitive clone (T. congolense IL 1180) or a clone expressing a moderate level of resistance to isometamidium (T. congolense IL 3343). Untreated control cattle were used to confirm the infectivity of each challenge. Of ten drug-treated cattle that were challenged with T. congolense IL 3343, all were refractory to infection at the first challenge. 1 month after drug administration. However, all ten animals succumbed to infection at either the second (seven cattle) or third (three cattle) monthly challenges. By contrast, all five drug-treated cattle challenged with T. congolense IL 1180 resisted four monthly challenges. The mean isometamidium concentration at the time of the first, 1 month, challenge was 5.6 +/- 2.8 ng/ml. At the time of the second monthly challenge the mean concentration was 2.0 +/- 0.86 ng/ml: at this time, concentrations were not significantly different between those cattle refractory to challenge with T. congolense IL 3343 and those cattle that were not. Thus, differences in susceptibility to challenge at this time would appear to be due to differences in the drug sensitivity of the parasite challenge. Finally, the mean isometamidium concentration in uninfected cattle at the time of the fourth monthly challenge was 0.4 +/- 0.18 ng/ml. These results indicate that when T. congolense infection occurs in cattle under isometamidium prophylaxis, the parasites may be considered at least moderately drug resistant if the concentration of isometamidium in serum is 2.0 ng/ml. At concentrations between 0.4 and 2.0 ng/ml a low level of drug resistance may be inferred. Below 0.4 ng/ml, however, no inference regarding drug resistance should be made.

Isometamidium concentrations in the sera of Boran cattle: correlation with prophylaxis against tsetse-transmitted Trypanosoma congolense.

Fifteen Boran cattle from a trypanosomiasis-free area were injected intramuscularly with isometamidium chloride at a dose of 1 mg/kg body weight. Thereafter, the cattle were challenged at monthly intervals with Glossina morsitans centralis infected with one of three populations of Trypanosoma congolense (IL 3893, IL 3889 or IL 1180) until all animals became infected. Isometamidium concentrations in the sera of these cattle were measured using a competitive enzyme-linked immunosorbent assay over the first 105 days following treatment. All cattle challenged with IL 3893 or IL 3889 developed infection following the first challenge, at which time the mean serum drug concentration in all treated cattle was 6 ng/ml. Cattle challenged with IL 1180 became infected following 6 to 8 monthly challenges. The mean serum drug concentration in these cattle at the time of their third monthly challenge with IL 1180 was 0.75 ng/ml. Trypanosome populations IL 3893 and IL 3889 were considered to be highly resistant to isometamidium, while IL 1180, relatively sensitive. It was therefore concluded that T. congolense persisting at serum isometamidium concentrations greater than 0.75 ng/ml can be considered moderately resistant, while those persisting at concentrations greater than 6 ng/ml can be considered markedly resistant. These results will be most valuable in the investigation of isometamidium resistance of T. congolense in the field.

The incidence of new and the prevalence and persistence of recurrent trypanosome infections in cattle in southwest Ethiopia exposed to a high challenge with drug-resistant parasites.

A method is described for calculating new infection incidence from monthly field data collected between April 1994 and February 1998 from an average of 770 Ethiopian Highland Zebu cattle maintained under traditional management in the Ghibe valley, southwest Ethiopia and exposed to a high challenge with drug-resistant parasites. Each month cattle with a packed cell volume (PCV)<26% and detected as parasitaemic, or cattle showing clinical signs of trypanosomosis, were treated with diminazene aceturate at 3.5 mg/kg body weight. An infection was defined as a new infection if it was preceded by 2 previous months in which both samples had a PCV>/=26% and were not detected with trypanosomes. Using this definition the average monthly incidence of infections of Trypanosoma congolense in cattle over 36 months of age was 13.3%, and the prevalence of recurrent infections 13.0%. Assuming that an animal had recovered from infection when PCV again returned to 26% without parasites being detected, mean persistence of infection was 3.8 months (median between 2 and 3 months). In contrast, T. vivax infections were susceptible to diminazene. The incidence of T. vivax infections in adults was lower than for T. congolense (2.8%), and they were less pathogenic (mean reduction of 3.1% units of PCV due to T. vivax infection compared with 4.6% units for T. congolense). Also, fewer cases were treated. Calves were first detected parasitaemic at an average age of 8.8 months and their infections persisted longer than those in adults. The effect of age on incidence of infection was not significant beyond 15 months of age. Adult male cattle appeared to be more susceptible to T. congolense infection than adult female cattle.