Modelling transmission of Mycobacterium avium subspecies paratuberculosis between Irish dairy cattle herds.

Bovine paratuberculosis is an endemic disease caused by Mycobacterium avium subspecies paratuberculosis (Map). Map is mainly transmitted between herds through movement of infected but undetected animals. Our objective was to investigate the effect of observed herd characteristics on Map spread on a national scale in Ireland. Herd characteristics included herd size, number of breeding bulls introduced, number of animals purchased and sold, and number of herds the focal herd purchases from and sells to. We used these characteristics to classify herds in accordance with their probability of becoming infected and of spreading infection to other herds. A stochastic individual-based model was used to represent herd demography and Map infection dynamics of each dairy cattle herd in Ireland. Data on herd size and composition, as well as birth, death, and culling events were used to characterize herd demography. Herds were connected with each other through observed animal trade movements. Data consisted of 13 353 herds, with 4 494 768 dairy female animals, and 72 991 breeding bulls. We showed that the probability of an infected animal being introduced into the herd increases both with an increasing number of animals that enter a herd via trade and number of herds from which animals are sourced. Herds that both buy and sell a lot of animals pose the highest infection risk to other herds and could therefore play an important role in Map spread between herds.

Quantifying transmission of Mycobacterium avium subsp. paratuberculosis among group-housed dairy calves.

Johne's disease (JD) is a chronic enteritis caused by Mycobacterium avium subsp. paratuberculosis (MAP), with control primarily aimed at preventing new infections among calves. The aim of the current study was to quantify calf-to-calf transmission of MAP among penmates in an experimental trial. Newborn Holstein bull calves (n = 32) were allocated into pens of 4, with 2 inoculated (IN) calves and 2 calves that were contact exposed (CE). Calves were group-housed for 3 months, with frequent collection of fecal and blood samples and tissue collection after euthanasia. The basic reproduction ratio (R0) was estimated using a final size (FS) model with a susceptible-infected model, based on INF-γ ELISA and tissue culture followed by qPCR. In addition, the transmission rate parameter (ß) for new shedding events was estimated using a general linearized method (GLM) model with a susceptible-infected-susceptible model based on culture, followed by qPCR, of fecal samples collected during group housing. The R0 was derived for IN and CE calves separately, due to a difference in susceptibility, as well as differences in duration of shedding events. Based on the FS model, interferon-γ results from blood samples resulted in a R 0 IG of 0.90 (0.24, 2.59) and tissue culture resulted in a R 0 T of 1.36 (0.45, 3.94). Based on the GLM model, the R0 for CE calves to begin shedding (R 0 CE ) was 3.24 (1.14, 7.41). We concluded that transmission of MAP infection between penmates occurred and that transmission among calves may be an important cause of persistent MAP infection on dairy farms that is currently uncontrolled for in current JD control programs.

Fecal shedding and tissue infections demonstrate transmission of Mycobacterium avium subsp. paratuberculosis in group-housed dairy calves.

Current Johne's disease control programs primarily focus on decreasing transmission of Mycobacterium avium subsp. paratuberculosis (MAP) from infectious adult cows to susceptible calves. However, potential transmission between calves is largely overlooked. The objective was to determine the extent of MAP infection in calves contact-exposed to infectious penmates. Thirty-two newborn Holstein-Friesian calves were grouped into 7 experimental groups of 4, consisting of 2 inoculated (IN) calves, and 2 contact-exposed (CE) calves, and 1 control pen with 4 non-exposed calves. Calves were group housed for 3 months, with fecal samples were collected 3 times per week, blood and environmental samples weekly, and tissue samples at the end of the trial. The IN calves exited the trial after 3 months of group housing, whereas CE calves were individually housed for an additional 3 months before euthanasia. Control calves were group-housed for the entire trial. All CE and IN calves had MAP-positive fecal samples during the period of group housing; however, fecal shedding had ceased at time of individual housing. All IN calves had MAP-positive tissue samples at necropsy, and 7 (50%) of the CE had positive tissue samples. None of the calves had a humoral immune response, whereas INF-γ responses were detected in all IN calves and 5 (36%) CE calves. In conclusion, new MAP infections occurred due to exposure of infectious penmates to contact calves. Therefore, calf-to-calf transmission is a potential route of uncontrolled transmission on cattle farms.

Epidemiological and economic consequences of purchasing livestock infected with Mycobacterium avium subsp. paratuberculosis.

BACKGROUND: Paratuberculosis (PTB) is a chronic disease which may lead to reduced milk yield, lower animal welfare and death in cattle. The causative agent is Mycobacterium avium subsp. paratuberculosis (MAP). The economic consequences are particularly important incentives in the control and eradication of the infection. One strategy to control PTB in a herd is to purchase animals from farms with a low risk of MAP infection. We wanted to investigate the epidemiological and economic consequences of buying livestock from different supplier farms of low, medium or high risk, as well as farms with unknown status. We also wanted to estimate the probability of spontaneous fadeout if the farmer of an initially MAP-free herd bought a specified number of infected animals in a single year, or continually bought infected animals. This was achieved through simulation modeling, and the effects of consistently introducing one, five or ten infected animals annually into an initially infection-free herd was also modeled. RESULTS: Our findings show that once infected, a farm can relatively safely purchase animals from other low and medium-risk farms without experiencing an increase in the prevalence, highlighting the importance of certification programmes. Furthermore, farms free of MAP are highly susceptible and cannot purchase more than a small number of animals per year without having a high risk of being infected. The probability of spontaneous fadeout after 10 years was 82% when introducing a single infected animal into an initially MAP-free herd. When purchasing ten infected animals, this probability was 46%. The continual purchase of infected animals resulted in very low probabilities of spontaneous fadeout. CONCLUSIONS: We demonstrated that MAP-free farms can purchase a small number of animals, preferably from certified farms, each year and still remain free of MAP. Already infected farms have little risk of increasing the prevalence on a farm when purchasing animals from other farms.

Presence of Mycobacterium avium subs. paratuberculosis DNA in milk used to feed calves in Portugal.

This Technical Research communication describes results of a study aimed at detecting the presence of Map in milk fed to calves, and identifying possible risk factors for that presence. A questionnaire was performed on 37 dairy farms and waste milk samples were collected on 3 occasions separated by a minimum of 1 week. For farms not feeding waste milk, bulk tank milk samples were collected instead. A real time PCR for the detection of the IS900 sequence was performed for the detection of Map. A majority of farms (89·2%) fed waste milk, with only one pasteurising the milk before feeding it to calves. Results of the PCR showed that 51·5% of the farms that were feeding waste milk had a positive result for Map on that milk. None of the studied risk factors were significantly associated with the presence of Map in milk samples, possibly due to the small number of farms entering the study. However, the prevalence of positive samples for Map on PCR was 3·5 times higher for farms that bought in animals from a single origin and 1·9 times higher for farms that bought from multiple farms, when compared with closed farms. Having a calving area for multiple cows also increased the risk of a positive Map result by 1·5 when compared with single pens. The risk of having a positive Map result on waste milk was 1·6 times higher for farms feeding that milk to male calves and 1·4 for farms feeding to both male and female calves, when compared with farms not feeding waste milk. This study highlights paratuberculosis as one of the potential risks of feeding waste milk to calves, and the need for mitigation strategies to be in place to avoid unnecessary disease transmission.

Modeling of Mycobacterium avium subsp. paratuberculosis dynamics in a dairy herd: An individual based approach.

In the dairy industry, Johne's disease (JD), caused by Mycobacterium avium subsp. paratuberculosis (MAP) is one of the major investigated diseases. To date, researchers have suggested some control strategies for JD, such as test-and-cull based herd management, isolated calf rearing management, and vaccinations. Due to the slow progressing nature of MAP, tests with low diagnostic test sensitivity and specificity, and economic limitations, implementing these strategies has not resulted in elimination of MAP from farms. To date, no study has integrated detailed dairy herd dynamics with different MAP transmission routes. We have developed an individual-based dairy herd model by incorporating basic herd dynamics in a closed herd environment where no new animals have been bought from outside. The model considered three age groups of animals: calves, heifers and adults. It includes sequential life events of a dairy animal and such key dynamic processes of the dairy herd as lactation cycle, calving, voluntary waiting period, insemination, pregnancy, dry-off period and calf and heifer rearing. After initially validating that the model reproduced typical herd dynamics, it was extended by incorporating MAP infection dynamics, where each individual adult animal belonged to one of four infection compartments: susceptible, latent, low shedding and high shedding. The model includes two disease transmission routes: horizontal transmission (i.e., fecal-oral) and vertical transmission (i.e., in utero infection). The results confirm that this model can simulate a realistic dairy herd and that inclusion of the above-mentioned dynamic processes provides useful information about individual infected animals to farmers. Access to the individual animal information offers more validity to assessment of appropriate control strategies for an endemically MAP infected herd. This model can serve as an accurate and novel tool not only to better understand MAP dynamics, but is also valuable as an individual based system of a typical dairy herd that can be applied to other research questions.

Impact of the shedding level on transmission of persistent infections in Mycobacterium avium subspecies paratuberculosis (MAP).

Super-shedders are infectious individuals that contribute a disproportionate amount of infectious pathogen load to the environment. A super-shedder host may produce up to 10,000 times more pathogens than other infectious hosts. Super-shedders have been reported for multiple human and animal diseases. If their contribution to infection dynamics was linear to the pathogen load, they would dominate infection dynamics. We here focus on quantifying the effect of super-shedders on the spread of infection in natural environments to test if such an effect actually occurs in Mycobacterium avium subspecies paratuberculosis (MAP). We study a case where the infection dynamics and the bacterial load shed by each host at every point in time are known. Using a maximum likelihood approach, we estimate the parameters of a model with multiple transmission routes, including direct contact, indirect contact and a background infection risk. We use longitudinal data from persistent infections (MAP), where infectious individuals have a wide distribution of infectious loads, ranging upward of three orders of magnitude. We show based on these parameters that the effect of super-shedders for MAP is limited and that the effect of the individual bacterial load is limited and the relationship between bacterial load and the infectiousness is highly concave. A 1000-fold increase in the bacterial contribution is equivalent to up to a 2-3 fold increase in infectiousness.

Environmental risk factors in the incidence of Johne's disease.

This review addresses the survival and persistence of Mycobacterium avium subsp. paratuberculosis (MAP), the causative pathogen of Johne's disease (JD), once it has left its ruminant host. JD has significant economic impact on dairy, beef and sheep industries and is difficult to control due to the long-term sub-clinical nature of the infection, intermittent or persistent MAP shedding during and after this period, inadequate test effectiveness, and the potential for MAP to exist for extended periods outside the host. The role that environmental factors play in the persistence and spread of MAP and consequent disease is assessed. Published risk factor analysis, organism survival across various environmental media and conditions, presence and spread in ruminant and non-ruminant wildlife, and the general potential for survival and multiplication of MAP ex-host both on and off-farm are discussed and knowledge gaps highlighted. An inclusive approach to disease management that takes into account the persistence and transport of the causative organism in on-farm soils and waters, land use and management, dispersal by domestic and non-domestic host species, as well as general animal husbandry is required on those farms where more traditional approaches to disease management have failed to reduce disease prevalence.

Modelling of paratuberculosis spread between dairy cattle farms at a regional scale.

Mycobacterium avium subsp. paratuberculosis (Map) causes Johne's disease, with large economic consequences for dairy cattle producers worldwide. Map spread between farms is mainly due to animal movements. Locally, herd size and management are expected to influence infection dynamics. To provide a better understanding of Map spread between dairy cattle farms at a regional scale, we describe the first spatio-temporal model accounting simultaneously for population and infection dynamics and indirect local transmission within dairy farms, and between-farm transmission through animal trade. This model is applied to Brittany, a French region characterized by a high density of dairy cattle, based on data on animal trade, herd size and farm management (birth, death, renewal, and culling) from 2005 to 2013 for 12,857 dairy farms. In all simulated scenarios, Map infection highly persisted at the metapopulation scale. The characteristics of initially infected farms strongly impacted the regional Map spread. Network-related features of incident farms influenced their ability to contaminate disease-free farms. At the herd level, we highlighted a balanced effect of the number of animals purchased: when large, it led to a high probability of farm infection but to a low persistence. This effect was reduced when prevalence in initially infected farms increased. Implications of our findings in the current enzootic situation are that the risk of infection quickly becomes high for farms buying more than three animals per year. Even in regions with a low proportion of infected farms, Map spread will not fade out spontaneously without the use of effective control strategies.

An immuno-epidemiological model for Johne's disease in cattle.

To better understand the mechanisms involved in the dynamics of Johne's disease in dairy cattle, this paper illustrates a novel way to link a within-host model for Mycobacterium avium ssp. paratuberculosis with an epidemiological model. The underlying variable in the within-host model is the time since infection. Two compartments, infected macrophages and T cells, of the within-host model feed into the epidemiological model through the direct transmission rate, disease-induced mortality rate, the vertical transmission rate, and the shedding of MAP into the environment. The epidemiological reproduction number depends on the within-host bacteria load in a complex way, exhibiting multiple peaks. A possible mechanism to account for the switch in shedding patterns of the bacteria in this disease is included in the within-host model, and its effect can be seen in the epidemiological reproduction model.

Dam Mycobacterium avium subspecies paratuberculosis (MAP) infection status does not predetermine calves for future shedding when raised in a contaminated environment: a cohort study.

Uptake of Mycobacterium avium subsp. paratuberculosis (MAP) by calves in the first days of life from colostrum, milk and faeces is regarded an important moment of transmission. The objective of this study was to quantify the association between the MAP status of dams as determined by the presence of MAP DNA and antibody in colostrum and that of DNA in faeces and the environment with subsequent MAP shedding of their daughters. A cohort of 117 dam-daughter pairs giving birth/being born on eight commercial dairy farms with endemic paratuberculosis was followed where colostrum, faecal and environmental samples (dust) were analysed for the presence of MAP using an IS900 real-time PCR. Antibodies in colostrum were measured by ELISA. Analysis of dust samples showed that on all farms environmental MAP exposure occurred continuously. In significantly more colostrum samples (48%) MAP DNA was detected compared to faecal samples (37%). MAP specific antibodies were present in 34% of the colostrum samples. In total MAP DNA was present in faecal samples of 41% of the daughters at least once during the sampling period. The association between faecal shedding in the offspring and the dam MAP status defined by MAP PCR on colostrum, MAP PCR on faeces or ELISA on colostrum was determined by an exact cox regression analysis for discrete data. The model indicated that the hazard for faecal shedding in daughters born to MAP positive dams was not significantly different compared to daughters born to MAP negative dams. When born to a dam with DNA positive faeces the HR was 1.05 (CI 0.6; 1.8) and with DNA positive colostrum the HR was 1.17 (CI 0.6; 2.3). When dam status was defined by a combination of both PCR outcomes (faeces and colostrum) and the ELISA outcome the HR was 1.26 (CI 0.9; 1.9). Therefore, this study indicates that neither the presence of MAP DNA in colostrum, MAP DNA in faeces nor the presence of MAP antibodies in colostrum of the dam significantly influences the hazard of MAP shedding in their subsequent daughters up to the age of two years when raised in a contaminated environment.

Factors affecting management changes on farms participating in a Johne's disease control program.

Modern Johne's disease programs aim to control Mycobacterium avium ssp. paratuberculosis (MAP) infection through implementation of management practices that reduce the probability of MAP introduction and within-herd transmission on dairy farms. Success of these programs depends on whether weaknesses in management are corrected through implementation of management improvements. The objectives of this study were, therefore, to (1) assess whether scores in risk-assessment (RA) questions predicted suggestions for management changes for the upcoming year; and (2) determine factors as assessed in an RA that motivated producers to make management improvements and assess whether management improvements were influenced by previously received test results. The RA determining on-farm management related to MAP introduction and transmission were conducted annually by herd veterinarians on 370 dairy farms participating in the Alberta Johne's Disease Initiative. A maximum of 3 management changes that the farmer and the veterinarian agreed upon were recorded in a management plan. The MAP infection status of the herds was assessed through culture of 6 environmental samples. Whereas a management change was proposed for only 4% of questions with scores 1 or 2 (low risk), a change was proposed for 19% of questions with scores >2 [high risk; odds ratio (OR)=11.4]. Improvement in RA question scores was more likely between the first and second annual RA than between the second and third RA (OR=1.6). Farms with >3 culture-positive environmental samples collected in the previous year were more likely to improve their management than environmental sample culture-negative farms (OR=1.3). In conclusion, proposed management changes were oriented toward previously identified weaknesses in management practices, suggesting that the RA was properly used to design targeted management suggestions. Furthermore, improvements in management were not randomly distributed among farms participating in the control program. Instead, knowledge of MAP infection status of a herd, suggestions for management improvements, and duration of participation all influenced implementation of management improvements.

Facts, myths and hypotheses on the zoonotic nature of Mycobacterium avium subspecies paratuberculosis.

Mycobacterium avium subspecies paratuberculosis (MAP) is the causative agent of paratuberculosis (Johne's disease [JD]), a chronic granulomatous enteritis in ruminants. JD is one of the most widespread bacterial diseases of domestic animals with significant economic impact. The histopathological picture of JD resembles that of Crohn's disease (CD), a human chronic inflammatory bowel disease of still unresolved aetiology. An aetiological relevance of MAP for CD has been proposed. This and the ambiguity of other published epidemiological findings raise the question whether MAP represents a zoonotic agent. In this review, we will discuss evidence that MAP has zoonotic capacity.

Shedding patterns of dairy calves experimentally infected with Mycobacterium avium subspecies paratuberculosis.

Although substantial fecal shedding is expected to start years after initial infection with Mycobacterium avium subspecies paratuberculosis (MAP), the potential for shedding by calves and therefore calf-to-calf transmission is underestimated in current Johne's disease (JD) control programs. Shedding patterns were determined in this study in experimentally infected calves. Fifty calves were challenged at 2 weeks or at 3, 6, 9 or 12 months of age (6 calves served as a control group). In each age group, 5 calves were inoculated with a low and 5 with a high dose of MAP. Fecal culture was performed monthly until necropsy at 17 months of age. Overall, 61% of inoculated calves, representing all age and dose groups, shed MAP in their feces at least once during the follow-up period. Although most calves shed sporadically, 4 calves in the 2-week and 3-month high dose groups shed at every sampling. In general, shedding peaked 2 months after inoculation. Calves inoculated at 2 weeks or 3 months with a high dose of MAP shed more frequently than those inoculated with a low dose. Calves shedding frequently had more culture-positive tissue locations and more severe gross and histological lesions at necropsy. In conclusion, calves inoculated up to 1 year of age shed MAP in their feces shortly after inoculation. Consequently, there is potential for MAP transfer between calves (especially if they are group housed) and therefore, JD control programs should consider young calves as a source of infection.

From mouth to macrophage: mechanisms of innate immune subversion by Mycobacterium avium subsp. paratuberculosis.

Johne's disease (JD) is a chronic enteric infection of cattle caused by Mycobacterium avium subsp. paratuberculosis (MAP). The high economic cost and potential zoonotic threat of JD have driven efforts to develop tools and approaches to effectively manage this disease within livestock herds. Efforts to control JD through traditional animal management practices are complicated by MAP's ability to cause long-term environmental contamination as well as difficulties associated with diagnosis of JD in the pre-clinical stages. As such, there is particular emphasis on the development of an effective vaccine. This is a daunting challenge, in large part due to MAP's ability to subvert protective host immune responses. Accordingly, there is a priority to understand MAP's interaction with the bovine host: this may inform rational targets and approaches for therapeutic intervention. Here we review the early host defenses encountered by MAP and the strategies employed by the pathogen to avert or subvert these responses, during the critical period between ingestion and the establishment of persistent infection in macrophages.

Pathogens at the livestock-wildlife interface in Western Alberta: does transmission route matter?

In southwestern Alberta, interactions between beef cattle and free-ranging elk (Cervus elaphus) may provide opportunities for pathogen transmission. To assess the importance of the transmission route on the potential for interspecies transmission, we conducted a cross-sectional study on four endemic livestock pathogens with three different transmission routes: Bovine Viral Diarrhea Virus and Bovine Herpesvirus 1 (predominantly direct transmission), Mycobacterium avium subsp. paratuberculosis (MAP) (indirect fecal-oral transmission), Neospora caninum (indirect transmission with definitive host). We assessed the occurrence of these pathogens in 28 cow-calf operations exposed or non-exposed to elk, and in 10 elk herds exposed or not to cattle. We characterized the effect of species commingling as a risk factor of pathogen exposure and documented the perceived risk of pathogen transmission at this wildlife-livestock interface in the rural community. Herpesviruses found in elk were elk-specific gamma-herpesviruses unrelated to cattle viruses. Pestivirus exposure in elk could not be ascertained to be of livestock origin. Evidence of MAP circulation was found in both elk and cattle, but there was no statistical effect of the species commingling. Finally, N. caninum was more frequently detected in elk exposed to cattle and this association was still significant after adjustment for herd and sampling year clustering, and individual elk age and sex. Only indirectly transmitted pathogens co-occurred in cattle and elk, indicating the potential importance of the transmission route in assessing the risk of pathogen transmission in multi-species grazing systems.

Spread of Mycobacterium avium subsp. paratuberculosis through soil and grass on a mouflon (Ovis aries) pasture.

The aims of this study were to describe spatial contamination of the environment on a mouflon pasture, as well as to assess the contamination of grass and roots after surface contamination and in depth contamination with feces and buried tissues from animals infected with Mycobacterium avium subsp. paratuberculosis (M. a. paratuberculosis). Samples of soil, roots, and aerial parts of plants were collected from different locations inside the mouflon pasture, and one control sample site was chosen outside the area where the animals are living. M. a. paratuberculosis DNA was present in all the examined sites and was more often detected in roots than in soil. DNA was detected at up to 80 cm of depth and was spatially more widespread than the initial hypothesis of M. a. paratuberculosis leaching vertically into deeper layers of soil. This study broadens our knowledge of the spread and persistence of M. a. paratuberculosis in an environment with highly infected animals.

Designing a risk-based surveillance program for Mycobacterium avium ssp. paratuberculosis in Norwegian dairy herds using multivariate statistical process control analysis.

Surveillance programs for animal diseases are critical to early disease detection and risk estimation and to documenting a population's disease status at a given time. The aim of this study was to describe a risk-based surveillance program for detecting Mycobacterium avium ssp. paratuberculosis (MAP) infection in Norwegian dairy cattle. The included risk factors for detecting MAP were purchase of cattle, combined cattle and goat farming, and location of the cattle farm in counties containing goats with MAP. The risk indicators included production data [culling of animals >3 yr of age, carcass conformation of animals >3 yr of age, milk production decrease in older lactating cows (lactations 3, 4, and 5)], and clinical data (diarrhea, enteritis, or both, in animals >3 yr of age). Except for combined cattle and goat farming and cattle farm location, all data were collected at the cow level and summarized at the herd level. Predefined risk factors and risk indicators were extracted from different national databases and combined in a multivariate statistical process control to obtain a risk assessment for each herd. The ordinary Hotelling's T(2) statistic was applied as a multivariate, standardized measure of difference between the current observed state and the average state of the risk factors for a given herd. To make the analysis more robust and adapt it to the slowly developing nature of MAP, monthly risk calculations were based on data accumulated during a 24-mo period. Monitoring of these variables was performed to identify outliers that may indicate deviance in one or more of the underlying processes. The highest-ranked herds were scattered all over Norway and clustered in high-density dairy cattle farm areas. The resulting rankings of herds are being used in the national surveillance program for MAP in 2014 to increase the sensitivity of the ongoing surveillance program in which 5 fecal samples for bacteriological examination are collected from 25 dairy herds. The use of multivariate statistical process control for selection of herds will be beneficial when a diagnostic test suitable for mass screening is available and validated on the Norwegian cattle population, thus making it possible to increase the number of sampled herds.

Meta-analysis of Johne's disease in the Iranian animal population (1999-2020).

Johne's disease (JD) affects domestic and wild animals across the globe. Paratuberculosis exerts huge economic impacts on the animal industry. Despite significant economic losses, little knowledge is available on the epidemiological status of Paratuberculosis in the animal population of Iran. The present study aimed to evaluate the prevalence rate of this disease in the Iranian animal population with confidence interval (CI) and p-value. The search was conducted on and screened the electronic international and national databases. Thereafter, sufficient and relevant data were extracted. Data were analyzed in STATA software (version 14). Prevalence disease rates were determined using random effect models. A total of 52 articles were included in the systematic review. According to the results, the overall disease incidence rate in Iran was 20.39%. The prevalence rate of JD was 22.33% (95% CI, 18.87-25.78) in the cattle population and 25.61% (95% CI, 21.43-29.78) in sheep. This study pinpointed that cattle and sheep were the most commonly infected hosts. The highest prevalence rate of disease was 35.88% in Tehran (95% CI, 16.77-54.99), followed by 32.86% (95% CI, 25.07-40.65), and 20.10% (95% CI, 14.63-25.58) in Khorasan Razavi and Kerman, respectively. The lowest prevalence rate of JD was 2.27% in Ilam (95% CI, 0.84-3.70). Based on this result, molecular-based methods were properly compared to other diagnostic methods. This study reported Mycobacterium avium subsp. paratuberculosis (MAP) prevalence in dairy herds in the provinces of Iran. The infection transmission from animal sources to humans and the potential role of MAP in human disease highlight a critical need for further study on this issue.

Suspicion of Mycobacterium avium subsp. paratuberculosis transmission between cattle and wild-living red deer (Cervus elaphus) by multitarget genotyping.

Multitarget genotyping of the etiologic agent Mycobacterium avium subsp. paratuberculosis is necessary for epidemiological tracing of paratuberculosis (Johne's disease). The study was undertaken to assess the informative value of different typing techniques and individual genome markers by investigation of M. avium subsp. paratuberculosis transmission between wild-living red deer and farmed cattle with known shared habitats. Fifty-three M. avium subsp. paratuberculosis type II isolates were differentiated by short sequence repeat analysis (SSR; 4 loci), mycobacterial interspersed repetitive-unit-variable-number tandem-repeat analysis (MIRU-VNTR; 8 loci), and restriction fragment length polymorphism analysis based on IS900 (IS900-RFLP) using BstEII and PstI digestion. Isolates originated from free-living red deer (Cervus elaphus) from Eifel National Park (n = 13), six cattle herds living in the area of this park (n = 23), and five cattle herds without any contact with these red deer (n = 17). Data based on individual herds and genotypes verified that SSR G2 repeats did not exhibit sufficient stability for epidemiological studies. Two common SSR profiles (without G2 repeats), nine MIRU-VNTR patterns, and nine IS900-RFLP patterns were detected, resulting in 17 genotypes when combined. A high genetic variability was found for red deer and cattle isolates within and outside Eifel National Park, but it was revealed only by combination of different typing techniques. Results imply that within this restricted area, wild-living and farmed animals maintain a reservoir for specific M. avium subsp. paratuberculosis genotypes. No host relation of genotypes was obtained. Results suggested that four genotypes had been transmitted between and within species and that one genotype had been transmitted between cattle herds only. Use of multitarget genotyping for M. avium subsp. paratuberculosis type II strains and sufficiently stable genetic markers is essential for reliable interpretations of epidemiological studies on paratuberculosis.