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Ruminant brucellosis in Upper Egypt (2005–2008)
Preventive Veterinary Medicine 101 (2011) 173–181
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Ruminant brucellosis in Upper Egypt (2005–2008) Y.M. Hegazy a,∗ , B. Molina-Flores b,d , H. Shafik c , A.L. Ridler e , F.J. Guitian f a
Department of Animal Medicine, Veterinary College, Kafrelsheikh University, Kafr Elsheikh, Egypt Egyptian Spanish Project for the Control of Brucellosis in the Upper Egypt, Spanish Agency for International Cooperation and Development, 21 El-Mahad El Swissry, Cairo, Egypt c General Organization for Veterinary Services, Ministry of Agriculture and Land Reclamation, Nadi El Said, Dokki 12311, Cairo, Egypt d Regional Animal Health Center for North Africa, Food and Agriculture Organization of the United Nations. 43, Avenue Kheireddine Pacha – 1002 Tunis, Tunisia e Institute of Veterinary, Animal & Biomedical Sciences IVABS, Tennent Drive, Massey University, 4442 New Zealand f Veterinary Epidemiology and Public Health Group, Department of Veterinary Clinical Sciences, The Royal Veterinary College, University of London, North Mymms, Hertfordshire AL9 7TA, UK b
a r t i c l e
i n f o
Article history: Received 25 March 2010 Received in revised form 9 April 2011 Accepted 11 May 2011 Keywords: Brucellosis Control program Households Ruminants Spatial analysis Upper Egypt
a b s t r a c t Brucellosis is endemic among humans and ruminant in Egypt and recent reports suggest that its incidence may be increasing. In this study we describe the frequency of brucellosis among different ruminant species in Upper Egypt and its spatial distribution using the data generated by a large-scale control campaign undertaken between 2005 and 2008. A total of 120,090 individual animals of different ruminant species were tested during the campaign. The true proportions of brucellosis were estimated as 0.79% (CI: 0.71%–0.87%), 0.13% (CI: 0.08%–0.18%), 1.16% (1.05%–1.27%) and 0.44% (0.34%–0.54%) among cattle, buffaloes, sheep and goats respectively. We estimated that 0.2% (CI: 0.16%–0.23%) of households in the study area keep at least one seropositive animal. Spatial autocorrelation of the proportions of seropositive households and seropositive animals was assessed using Global Univariate Moran’s I and Local Univariate LISA. These analyses showed that the distribution of seropositive animals has considerable spatial heterogeneity with clustering in the northern governorates of the study area. Our results show that brucellosis is widespread and heterogeneously distributed in Upper Egypt. At the current level of available resources it is very unlikely that test and slaughter could be implemented with the intensity needed to be effective and other control measures that could replace or complement the test and slaughter policy in place should be considered. Also, this study illustrates some of the challenges faced by bilateral projects that have to accommodate an externally funded intervention with an ongoing national official disease control program. © 2011 Elsevier B.V. All rights reserved.
1. Introduction Brucellosis is one of the world’s most widespread zoonotic diseases caused by species of the genus Brucella (WHO, 2009; Moreno et al., 2002). Infection in ruminants causes economic losses due to abortion, infertility and pre-
∗ Corresponding author. Tel.: +20 2402790113. E-mail address: [email protected] (Y.M. Hegazy). 0167-5877/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.prevetmed.2011.05.007
mature culling (Radostits et al., 2000). Control of brucellosis depends generally on test and isolation/slaughter of positive animals, vaccination of susceptible animals and control of animal movements (Corbel, 1997). Various countries all over the world have implemented different control policies using combinations of these control measures, with reported success in some countries and failure in others (Cutler et al., 2005; Blasco, 1997). In Egypt, since 1981, the General Organization of Veterinary Services (GOVS) has run a control program which
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is currently based on testing female ruminants older than 6 months of age with slaughter of serologically positives, and voluntary vaccination of calves using Brucella abortus S19 vaccine and Brucella melitensis Rev 1 vaccine for lambs and kids (Refai, 2002). Despite almost 30 years of implementation of the control program, brucellosis remains endemic among ruminants and humans in Egypt and recent reports suggest that the incidence of human infection may be increasing (Jennings et al., 2007). Given that infected animals are the source of human infection, the increasing incidence of human brucellosis probably reflects a similar trend in domestic animals. Unfortunately, reliable data on the frequency and distribution of ruminant brucellosis in Egypt is lacking. Perhaps a reason why the official control program has limited success and has not been always fully adhered to is that the strategies were not adequate for the baseline frequency of disease (Hegazy et al., 2009). Indeed, it is generally accepted that depending on the prevalence/incidence of brucellosis among different animal species, different combinations of control strategies may be appropriate (Robinson, 2003). A better understanding of the frequency of infection and its distribution across geographic areas may help tailor the control program to the situation and may contribute to both its actual implementation as per official guidelines and its impact in reducing the frequency of infection in livestock and humans. This is the overall aim of the present study, which we have tried to achieve for the Upper Egypt region, an area in the valley of the Nile river, south of the delta, which contains around 32% and 39% of the total Egyptian large and small ruminant population respectively (Ministry of Agriculture and Land Reclamation, 2005). In this area it is estimated that more than 90% of the large ruminants are kept by households owning between 1 and 5 cattle and/or buffaloes which are often in close contact with sheep and goats (Aidaros, 2005). Accurate and unbiased estimates of the frequency of brucellosis in the region are lacking, but a recent study in two of the Upper Egypt governorates found between 4.5 and 7% of seropositive animals among different ruminant species (Samaha et al., 2008). Lack of information on the target population and sampling procedure for villages/farms and animals within them make the results of this study difficult to extrapolate to the general population. However, if the results are not a gross overrepresentation of the individual-level true prevalence of infection, they seriously question the appropriateness of the current control strategy based on test and slaughter of positive animals and voluntary vaccination of young animals. Only if the prevalence was considerably lower and enough resources were available for frequent testing of a large enough proportion of the population would such a program be likely to succeed (Hegazy et al., 2009). In the current situation, estimates of the frequency of infection are urgently required to make a preliminary assessment of the appropriateness of the control strategy in place. In 2005, a 3-years long Egyptian-Spanish bilateral project for the control of brucellosis in Upper Egypt was initiated and jointly implemented by the GOVS and the Spanish Agency for International Cooperation and
Development (AECID). The project aimed at reducing the incidence of brucellosis in the ruminant population of Upper Egypt by providing support to the GOVS control program. Specifically, the proportion of adult ruminants tested in each governorate would be increased concurrently with a vaccination campaign. Assuming probabilistic selection of animals to be tested and the slaughter of those found positive it was expected that the enhanced control program would (i) generate better prevalence estimates (ii) reduce the incidence of ruminant brucellosis in the region. The primary objective of this study is to describe the frequency of brucellosis among different ruminant species in Upper Egypt and its spatial pattern using the results of the 2005–2008 brucellosis control campaign carried out in this area. A secondary objective is to describe this bilateral control program and in particular the main challenges and lessons learned from the implementation of a large scale brucellosis control program in an endemic area. 2. Materials and methods 2.1. Study area and control campaign activities The study area consisted of the 7 Upper Egypt governorates of Beni Sueif, Al Minia, Assuit, Sohag, Quena, Luxor and Aswan. The governorates of Al Fayoum and Al Giza as well as the southern area of Cairo are often included as part of Upper Egypt but for purpose of this study they are excluded. Within these 7 governorates only 6.4% of the land area is populated, the remainder is desert (Fig. 1). Our study was restricted to the populated area of the 7 governorates, which includes 55 administrative units (districts) with 430 veterinary units. Veterinary units are governmental units distributed all over Egypt (around 1500 as mentioned by Aidaros, 2005) managed by one or more official veterinarians. These units are responsible for supplying different veterinary services to the livestock sector including the application of mandatory interventions. The study area represents 13% of the total populated area of Egypt. The control campaign was initiated in October 2005 and finished in October 2008. It was planned for this campaign to consist of 2 main activities: - First, yearly serological testing against Brucella spp. of a randomly selected 10% of the total adult ruminant population in each governorate (130,999 total animals to be tested from the whole of Upper Egypt, yearly). The number of animals to be tested every year was calculated in order to allow yearly estimation of the prevalence of seropositive animals of the different species (cattle, buffalo, sheep, goats) in each governorate, with 90% confidence and 15% precision, using as expected prevalence values the % of seropositive animals among those tested as a result of the GOVS control activities in 2003, previous to the start of the project. The resulting number of animals of the different species to be sampled in each governorate was divided equally among the local veterinary units of the governorate. The veterinarian in charge of the veterinary unit was asked to select randomly the allocated number of samples and to collect the samples from all villages served by the clinic and from differ-
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2.2. Data sources The available data for this study consisted of information on the sex, age, breed, farm/household location (governorate, district and village), owner identification, date of sampling and results of the serological tests for all individual animals sampled as part of the control campaign between October 2005 and January 2008. This information was stored in a relational database that was developed as part of the campaign. Each veterinary unit was provided with a computer supplied with software specifically built by the project for recording of the required data for each animal. The veterinarians received training on how to collect the field data using predesigned forms and how to use the database and introduce data remotely. 2.3. Data analysis For those animals that were tested more than once (4113 animals), only the first record was used in the analysis. 2.4. Descriptive statistics Descriptive statistics were obtained for the key variables in relation to the different aspects of the campaign such as numbers of tested animals, villages and local veterinary units involved in the campaign. Fig. 1. Map of Upper Egypt showing the location of the seven governorates of the study area and their populated and desert areas.
ent farms within the village, however, random sampling was not fully adhered to. This was attributed to lack of incentives for veterinarians and poor cooperation from farmers. Also, no further explanation was provided with regard to what random selection involves and how it can be performed across different levels (selection of households or farms within villages and of individual animals within households or farms). This initial annual target was revised soon after the start of the campaign to the same number of animals to be tested but across all the 3 years of the campaign rather than annually. Such testing would be carried out while the GOVS would continue to be responsible for slaughtering of positive reactors and compensating the owners. Serological testing consisted of Rose Bengal Plate test (RBPT) followed by Complement Fixation test (CFT) for animals seropositive to the former test (series interpretation), as officially approved by the national veterinary services in Egypt in accordance with the OIE manual of standard diagnostic tests and vaccines (World Organization for Animal Health, 2004). - Second, mass vaccination of all female cattle and buffaloes with Brucella abortus S19 at a dose of 1010 and of all sheep and goats (males and females) with Brucella melitensis Rev1 at a dose of 109 .
These activities were to be undertaken by 964 veterinarians and 2396 assistants from 430 local veterinary units.
2.5. Seroprevalence of brucellosis at individual animal level The apparent prevalence was calculated as the number of animals found to be seropositive divided by the total number of animals sampled during the study period. It was obtained both, for all species combined and by species. The true proportion of seropositive animals among those tested (TP) and the true proportion of seropositive animals by species (STP) were calculated after adjusting for the combined sensitivity (CSe) and combined specificity (CSp) of the serological tests from the apparent prevalence (AP) as TP = (AP + CSp − 1)/(CSe + CSp − 1). The values of CSe and CSp of the testing strategy were obtained, assuming series interpretation, as the most likely values from triangular distributions for CSe = SeRBT × SeCF and for CSp = 1 − ((1 − SpRBT ) × (1 − SpCF )); where SeRBT , SpRBT, SeCF, SpCF are the sensitivity and specificity of RBPT and CFT respectively. A very wide range of values for Se and Sp are presented in the published literature; to account for this variability, the values of Se and Sp of individual tests were obtained by random selection from uniform distributions with minimum and maximum values derived from the literature: 0.72 < SeRBT < 1; 0.8 < SpRBT < 1; 0.81 < SeCF < 1; 0.80 < SpCF < 1 (Blasco et al., 1994; Gall and Nielsen, 2004; Ramirez-Pfeiffer et al., 2007). Ten-thousand values of the sensitivity and specificity of both serological tests were chosen from the uniform distributions to generate the distributions of CSe and CSp. Values of CSe and CSp were assumed to be the same across species. 95% confidence intervals (CI) around the estimated proportion were obtained as a measure of precision of the estimates
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(Newcombe, 1998). The STP was also calculated by year. The true proportion of seropositive animals by district (DTP) was estimated the same way as TP and STP. 2.6. Seroprevalence of brucellosis at household level For each district, the proportion of “tested households” with serologically positive animals (household-level district apparent prevalence: HDAP) was obtained. Any household for which one or more tested animals were found seropositive was considered a seropositive household. Information on herd/flock size for those households in which animals were sampled was not available but the vast majority of households keeping ruminants in this part of Egypt have just a few animals (Aidaros, 2005). It was assumed in this study that the total number of animals tested from a single owner was the total size of the household herd/flock – i.e. it was assumed that once a household was included in the campaign all its animals had been sampled. Using estimated values of householdlevel sensitivity and specificity (details below) the true proportion of seropositive households by district (HDTP) was estimated from the apparent proportion (HDAP) as HDTP = (HDAP + HCSp − 1)/(HCSe + HCSp − 1), where HCSp is the household-level combined specificity and HCSe the household level combined sensitivity. The apparent and true proportion of households with seropositive animals for brucellosis in each governorate of Upper Egypt (HGAP and HGTP) were calculated in the same way as HDTP. The apparent and the true overall prevalence (HAP and HTP) for the whole area of Upper Egypt were calculated in the same way. 2.7. Simulation of the likely performance of the testing strategy at household level The likely ability of the testing strategy to correctly identify households with and without infected animals was assessed by means of simulation. For purpose of this simulation it was assumed:
of infected households with seropositive animals, noninfected households with seropositive animals, infected households without seropositive animals and non-infected households without seropositive animals were used to calculate the HCSe and HCSp. 2.8. Spatial distribution of seropositive animals and households To visualize the geographic distribution of ruminant brucellosis in Upper Egypt, two choropleth maps representing the estimated true prevalence of seropositive animals and the estimated true prevalence of seropositive households per district were created using Arc GIS 9.2 (ESRI 2006). To assess the extent to which the status of a district with regard to brucellosis (as defined by the proportion of seropositive animals and the proportion of its households with seropositive animals) was spatially correlated within the study area, we calculated the global univariate Moran’s I considering any district that shares any common point with another district as neighbor (first order queen contiguity). Since some of the districts had few households and animals tested, numbers of animals and households sampled per district were used in an empirical Bayes smoothing method to adjust for underlying population structure (Henning et al., 2009). Inference for the significance of the value of Moran’s I obtained was assessed by running this analysis for 999 permutations. In order to identify areas with either homogeneously high or low seroprevalence of brucellosis we used Local Univariate LISA (Local Indicators of Spatial Association) to examine the local level of spatial autocorrelation. The same settings as for the Global Univariate Moran’s I were used; significant clusters were identified after running the analysis for 999 permutations at P < 0.05. These analyses were carried out using GeoDa 0.9.5-i.5 (http://geoda.uiuc.edu/). 3. Results 3.1. Proportion of seropositive animals
- All individual animals in the population were equally likely to be infected regardless of their aggregation within households (intracluster correlation coefficient () = 0). - All households were equally likely to be sampled with probability = 0.1. - The distribution of number of animals per household differed between districts. Household size was assumed to follow a triangular distribution with parameters varying between districts; minimum and maximum values where derived from the distribution of animals sampled from the same owner by district. When a household with n animals is sampled, any number of animals from 1 to n can be sampled with equal probability. - The combined sensitivity and specificity of the tests interpreted in series at individual animal level were used as calculated above. The simulation was carried out in @Risk software (version 3.5d, Palisade Corporation, Newfield, NY, USA). The simulation was run for 10,000 iterations. The total number
A total of 120,077 individual animals of different ruminant species were tested between October 2005 and January 2008 and had complete information. Twelve percent of the local veterinary units did not participate in animal sampling during this period, the remaining 88% veterinary units collected samples from 54% of the villages in the study area, which suggests some clustering of sampling by village (Table 1). The CSe and CSp of the series combination of the two tests ranged triangularly from 64% to 92% and 97% to 100% respectively with most likely values found to be 78% and 99–100% respectively. A total of 1071 animals were found seropositive to RBPT and 693 of them were found positive to CFT. The TP was estimated as 0.74% (CI: 0.69%–0.79%). By species, the STP were estimated as 0.79% (CI: 0.71%–0.87%), 0.13% (CI: 0.08%–0.18%), 1.16% (1.05%–1.27%) and 0.44% (0.34%–0.54%) for cattle, buffaloes, sheep and goats respectively. Only data from years 2006 and 2007 will be used in the analyses by year (the results of 2005 and 2008 were
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the proportion ranging from 0.1% to 3.0% in the different governorates and years of the campaign (Table 2). Based on our simulations, the HCSe and HCSp, ranged from 47.0% to 73.0% and from 95.0% to 97.0% respectively according to district. If our assumptions hold true, the overall HTP would be 0.2% (CI: 0.16%–0.23%). The HDTP were calculated and showed a large fluctuation (from 0.1% to 7.7%) between districts as shown in Fig. 3. The HGAP and HGTP for different governorates are shown in Table 2.
3.3. Spatial distribution of seropositive animals and households Fig. 2. Estimated true proportions of serologically positive individuals of different ruminant species against Brucella spp. in Upper Egypt 2006–2007.
omitted, as there were low numbers of animals and households sampled in this period; 21 animals in 2005 and 1859 animals in 2008). In 2006 the numbers of tested animals were 30,154 cattle, 6615 buffaloes, 19,046 sheep and 7252 goats, while for 2007, 19,004 cattle, 11,438 buffaloes, 13,987 sheep and 10,701 goats. The STP in 2006 and 2007 by species are presented in Fig. 2 and the overall DTP for the combined result of years 2006 and 2007 is shown in Fig. 3. 3.2. Seroprevalence at household level Overall, 1.2% (CI: 1.1%–1.3%) of the total households tested had seropositive animals against Brucella spp., with
Districts in the northern governorates of the study area (Beni sueif and Al Minia) had the highest proportions of positive animals. There was a tendency towards a declining proportion of seropositive animals from Northern to Southern districts in the study area (Fig. 3). The value of the Global Moran’s I using empirical Bayes smoothing for individual animal prevalence was 0.58 (P < 0.001) indicating that there was a significant spatial autocorrelation (clustering) of the proportion of positive individual animals between districts. The results of location of clusters identification using LISA with spatial empirical Bayes indicated that there was significant spatial clustering (P < 0.05) between districts with high proportions of seropositive animals in Beni Sueif and Al Minia governorates (northern part of the study area). On the other hand, there was a tendency towards increasing proportion of seropositive households from Northern to Southern districts in the study area (Fig. 3). The
Fig. 3. Distribution of estimated true proportion of individual ruminants (A) and households (B) seropositive against Brucella spp. among Upper Egypt districts 2006–2007.
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Fig. 4. LISA cluster maps for identification of districts with similar proportions of (A) ruminants seropositive against Brucella spp. and (B) households seropositive against Brucella spp. in Upper Egypt, 2006–2007.
value of the Global Moran’s I using empirical Bayes smoothing for household prevalence was – 0.013 (P < 0.445) which indicates a non significant clustering of seropositive households. Results of local cluster identification showed that there was clustering of districts with low proportions of seropositive households in Beni Sueif governorate (Fig. 4). 4. Discussion Brucellosis is an endemic disease of humans and ruminants throughout most of the Middle East and some countries of the Mediterranean basin (Refai, 2002; Benkirane, 2006; Pappas et al., 2006). Its economic and public health importance has lead countries in the area to establish different control programs (Refai, 2002). In Egypt, the GOVS has run a program for the surveillance and control of ruminant brucellosis since 1980. The main elements of this control program consist of serological testing every 6 months of all female cows, buffaloes, sheep and goats over 6 months old and valuable bulls, rams and bucks using RBPT and CFT with slaughtering of seropositive animals and compensation of the owners. This is accompanied by voluntary vaccination of calves, lambs and kids which are serologically negative at 3–6 months age, using S19 vaccine for cattle and buffaloes and Rev1 vaccine for sheep and goats (Refai, 2002). A recent study showed that no more than 7% of target animals were tested every year (Hegazy et al., 2009), because of lack of farmers and veterinarians incentives and insufficient funding to sustain the application of this policy. There are no accurate figures for the actual coverage of the official vaccination program, but from dis-
cussions with the project manager of the campaign and with veterinarians from the Egyptian GOVS and on the basis of figures previously reported to the OIE, the voluntary vaccination program appears to have very limited coverage and unlikely to result in the vaccination of more than 1% of the calves, lambs and kids. These factors have resulted in the inability of this program to decrease the prevalence of brucellosis in Egypt (Refai, 2002; Hegazy et al., 2009, 2011). It has also resulted in the absence of reliable estimates of the frequency of infection and how it varies across species, production systems and geographic areas. The main reason why such estimates are not available is lack of a formal sampling strategy; it has been suggested that, at least for some parts of the country, the implementation of the test and slaughter campaign relies on haphazard sampling concentrated in areas that are accessible and with huge variations in its intensity across species year to year (Hegazy et al., 2009). One of the objectives of the 2005–2008 control campaign was to decrease the prevalence of brucellosis among different ruminant species by means of obligatory whole herd vaccination. The other objective was to generate data that could be used to estimate the seroprevalence of brucellosis in Upper Egypt and, accordingly, the campaign aimed at probabilistic sampling of individual animals. In this study, we attempted to fulfill some of the knowledge gaps with regard to the frequency and distribution of brucellosis in Upper Egypt by analyzing the large amount of data generated by the 2005–2008 control campaign. Over 3 years (October 2005 to January 2008), the campaign succeeded in testing around 9% of the total adult
Table 1 Total numbers of local veterinary units in Upper Egypt and distribution of these veterinary units regarding the proportions of their served villages which are sampled for brucellosis as part of the Egyptian-Spanish bilateral project for the control of brucellosis in Upper Egypt in 2006 and 2007. Total number of Veterinary Units
Number of Veterinary Units which did not test
*
Veterinary Units which tested only one village
Veterinary Units which tested <50% of their associated villages
Veterinary Units which tested <75% of their associated villages
Total number of villages
Number of villages sampled
Beni Sueif Al Minia Assuit Sohag Quena Luxor Aswan Total
76 90 77 80 48 11 48 430
2 (3%) 3 (3%) 25 (32%) 14 (17%) 1 (2%) 0 (0%) 5 (10%) 50 (12%)
16 (21%) 8 (9%) 15 (19%) 16 (20%) 2 (4%) 0 (0%) 2 (4%) 59 (14%)
32 (42%) 26 (29%) 44 (57%) 42 (53%) 7 (15%) 1 (9%) 10 (21%) 162 (38%)
52 (68%) 54 (60%) 58 (75%) 64 (80%) 20 (42%) 3 (27%) 20 (42%) 271 (63%)
466 612 359 570 334 96 252 2689
227 (49%) 358 (58%) 138 (38%) 253 (44%) 243 (73%) 70 (73%) 158 (63%) 1447 (54%)
*
These Veterinary units serve more than one village.
Table 2 Results of serological testing against Brucella spp. of households containing ruminants as a part of the Egyptian-Spanish bilateral project for the control of ruminant brucellosis in Upper Egypt in 2006 and 2007. Year
2006
Governorate Total number of households tested Beni Sueif Al Minia Assuit Sohag Quena Luxor Aswan Total
6703 3868 2778 2658 4236 478 1748 22,469
2007 Number of negative households 6502 3822 2764 2651 4224 477 1747 22,187
Number and % of positive households 201 (3.0%) 46 (1.2%) 14 (0.5%) 7 (0.3%) 12 (0.3%) 1 (0.2%) 1 (0.1%) 282 (1.3%)
Total number of households tested 4428 3442 280 3578 3494 1684 909 17,822
Number of negative households 4294 3412 280 3585 3486 908 906 17,638
Number and % of positive households 134 (3.0%) 30 (0.9%) 0 (0.0%) 7 (0.2%) 8 (0.2%) 2 (0.1%) 3 (0.3%) 184 (1.0%)
Total number of tested households 11131 7310 3058 6243 7730 2162 2657 40,291
Apparent proportion of True proportion of positive households positive households %(HGAP) %(HGTP) (CI) 3.0% (2.7–3.3) 1.0% (0.8 –1.2) 0.5% (0.3–0.7) 0.2% (0.1–0.3) 0.3% (0.2–0.4) 0.1% (0.0–0.3) 0.2% (0.0–0.4) 1.2% (1.1–1.3)
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Governorate
0.3% (0.2–0.4) 0.1% (0.0–0.2) 0.4% (0.2–0.6) 0.2% (0.1–0.3) 0.1% (0.0–0.2) 0.1% (0.0–0.2) 0.2% (0.0–0.4) 0.2% (0.16–0.23)
CI, confidence interval.
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ruminant population in Upper Egypt with slaughtering of 125 seropositive animals. The remaining seropositive animals were not slaughtered because of the lack of available funding to compensate their owners. The estimated true proportions of seropositive animals in the study area ranged from 0.13% in buffaloes to 1.16% in sheep. This seroprevalence estimate should not have been substantially affected by the vaccination campaign, given the low coverage. However, caution should be taken when interpreting these estimates which may not reflect the actual proportion of positive animals in the study area because of some weaknesses: despite the instructions given to the local veterinarians that random selection should have been used, they were not given further explanations on what random selection actually implies and how it can be achieved. Furthermore, veterinarians were expected to carry out the sampling as part of their normal duties without additional incentives and no supervision or formal control of the way they conducted the sampling was in place. These limitations raise considerable doubts as to the extent to which probabilistic selection was achieved, limiting our ability to extrapolate the obtained results to the reference livestock population of Upper Egypt. We estimated that 0.2% of all households in the study area had seropositive animals. This figure accounts for the inadequate performance of diagnostic tests (based on our assumptions) but, similarly to the animal-level estimates, it may be affected by selection bias due to non-random selection of households and animals within households. However, we believe that household-level estimates are probably less influenced by departures from probabilistic sampling and aggregation of samples within certain clusters than individual animal estimates. Overestimation of the true proportion of positive households due to selection bias cannot be ruled out, but we consider this to be unlikely. Based on our experience in the area, selection bias would be due to overrepresentation of households nearby the veterinary clinics and therefore unrelated to brucellosis status. The main Brucella species responsible for infection in ruminants are Brucella abortus and Brucella melitensis. In this research, we did not identify the Brucella species in infected animals. However, the main isolate in all animal species and humans in Egypt is Brucella melitensis (Refai, 2002) and there is evidence that suggests that cattle act as spill-over hosts of this species (Hegazy et al., 2011). Our results suggest that ruminant brucellosis is not homogeneously distributed across the study area, with higher prevalence of infection in northern districts. A possible explanation may be the location of these districts closer to the large animal markets in Cairo where animals from different sources, including the Nile delta mix and are redistributed (Muma et al., 2007). A second factor that could contribute to the higher prevalence of infection observed in the northern governorates of Upper Egypt is that this area has one of the highest densities of ruminants in Egypt, in particular sheep and goats, which are often reared in movable flocks (Ministry of Agriculture and Land Reclamation, 2005). These types of flocks are particularly common in Al Minia governorate (Fayed, personal communication).
Prevalence estimates are the cornerstone for deciding the appropriate control policy for brucellosis and for testing its efficiency. Despite the vast amount of data it generated, the 2005–2008 campaign does not provide prevalence estimates that could assertively be regarded as unbiased. At the same time, the results obtained provide some insight on the likely disease distribution among different ruminant species and different geographic areas in Upper Egypt. As for the efficiency of the campaign in reducing the frequency of infection in the target population, only a small proportion of the ruminant population was tested every year. Testing such a small proportion of the population with slaughtering of positive animals would allow the infection to persist. Consequently, the cost-effectiveness of the campaign seems to be, at least, questionable. Furthermore, lack of funding for compensation of livestock owners did not allow for all animals found positive to be culled. The study illustrates some of the challenges resulting from the need to accommodate the activities of an externally funded 3-year intervention and an ongoing official disease control program. Soon after the project was started its objectives were revised. The original intention of achieving probabilistic sampling of 10% of the total ruminant population of Upper Egypt every year was soon replaced for the same proportion but during the whole 3-year duration of the program and the vaccination campaign had to be delayed until the very end of the project because of logistic difficulties (it is currently being undertaken). A major challenge of this particular control program has been that a sampling strategy aimed at gaining knowledge on the frequency of infection (as suggested by the intention of randomly selecting animals to be tested stated in the protocol of the campaign) had to be coordinated with the official test and slaughter control policy. Some of the insights into the situation of ruminant brucellosis in Upper Egypt gained from this study should be taken into account when designing the next phases of the official control campaign. The current test and slaughter policy is a reasonable strategy for the control of brucellosis in Upper Egypt only if our estimates for the proportion of seropositive animals and households are not heavily biased downwards and if a sufficient proportion of the ruminant population could be tested frequently enough (Corbel, 2006; Hegazy et al., 2009) accompanied with slaughtering of seropositive animals. Even with external assistance, it has proved challenging to test 10% of the ruminant population in a 3-year period, therefore it seems unlikely that with the current level of resources test and slaughter could be implemented with the intensity needed to significantly reduce the prevalence of infection among ruminants in the area. For that reason, the national control program should also consider other control strategies that can replace or complement test and slaughter such as compulsory vaccination and more strict application of quarantine and movement control (Minas, 2006). The observed spatial heterogeneity may offer opportunities for targeted interventions in the ruminant population. Interventions that could minimize the spread of the infection from the Northern governorates of Upper Egypt to non-infected areas or areas with lower prevalence may be a cost-effective strategy (Mainar-Jaime et al., 2005). Geographic zoning as a
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policy for brucellosis control and/or eradication has been successfully implemented in some countries such as Chile, where brucellosis eradication programs were initiated in regions with low cattle density and low prevalence, and then extended gradually to other regions (Rivera et al., 2002). Similar strategies could be particularly appropriate in countries with scarce economic resources such as Egypt. A possible alternative to the current control program could be compulsory vaccination in geographic areas with high seroprevalence (FAO/WHO/OIE, 1993). For such interventions to succeed in reducing the incidence of ruminant brucellosis additional measures such as control of animal movement would be required. Crucially, as the 2005–2008 campaign clearly shows, the success of any control strategy for brucellosis in Upper Egypt has to consider the perceptions and incentives of farmers and veterinarians and try to engage them. The spatial aggregation of districts with relatively high proportions of seropositive animals in the Northern governorates of Upper Egypt, suggests that the risk of human exposure to Brucella spp. maybe particularly high among rural households in this region. Public health services should probably pay particular attention to the rural population of this area during their campaigns for prevention of human brucellosis. 5. Conclusion Brucellosis is widespread among different ruminant species in Upper Egypt, where we estimate that 0.2% of households keep seropositive ruminants. Infection is heterogeneously distributed, with highest prevalence in the ruminant population of the Northern areas and as a result higher risk of human exposure. At the current level of available resources it is very unlikely that test and slaughter could be implemented with the intensity needed to be effective and other control measures should be considered. Acknowledgements We would like to acknowledge the staff members of the General Organization of Veterinary Services in Cairo and ˜ de Cooperación Internacional para el the Agencia Espanola Desarrollo for their help with the collection of data and the permission to use these data in this study. References Aidaros, H., 2005. Global perspectives – the Middle East: Egypt. Rev. Sci. Tech. Off. Int. Epiz. 24, 589–596. Benkirane, A., 2006. Ovine and caprine brucellosis: world distribution and control/eradication strategies in West Asia/North Africa region. Small Rum. Res. 62, 19–25. Blasco, J.M., 1997. A review of the use of B. melitensis Rev 1 vaccine in adult sheep and goats. Prev. Vet. Med. 31, 275–283.
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Blasco, J.M., Marín, C., Jiménez de Bagués, M., Barberan, M., Hernandez, A., Molina, L., Velasco, J., Diaz, R., Moriyon, I., 1994. Evaluation of allergic and serological tests for diagnosing Brucella melitensis infection in sheep. J. Clinic. Microbiol. 32, 1835–1840. Corbel, M.J., 1997. Brucellosis: an overview. Emerg. Infect. Dis. 3, 213–221. Corbel, M.J., 2006. Brucellosis in Humans and Animals. WHO Press, World Health Organization, Geneva. Cutler, S.J., Whatmore, A.M., Commander, N.J., 2005. A review brucellosis – new aspects of an old disease. J. Appl. Microbiol. 98, 1270–1281. FAO/WHO/OIE, 1993. FAO/WHO/OIE Guidelines for a Regional Brucellosis Control Programme for the Middle East. Amman, Jordan. Gall, D., Nielsen, K., 2004. Serological diagnosis of bovine brucellosis: a review of test performance and cost comparison. Rev. Sci. Tech. Off. Int. Epiz. 23, 989–1002. Hegazy, Y.M., Ridler, A.L., Guitian, F.J., 2009. Assessment and simulation of the implementation of brucellosis control programme in an endemic area of the Middle East. Epidemiol. Infect. 137, 1436–1448. Hegazy, Y.M., Moawad, A., Osman, S., Ridler, A., Guitian, J., 2011. Ruminant Brucellosis in the Kafr El Sheikh Governorate of the Nile Delta, Egypt: Prevalence of a Neglected Zoonosis. PLoS Negl. Trop. Dis. 5 (1), e944, doi:10.1371/journal.pntd.0000944. Henning, J., Pfeiffer, D.U., Le Tri, V.U., 2009. Risk factors and characteristics of H5N1 Highly Pathogenic Avian Influenza (HPAI) post-vaccination outbreaks. Vet. Res. 40, 15. Jennings, G.J., Hajjeh, R.A., Girgis, F.Y., Fadeel, M.A., Maksoud, M.A., Wasfy, M.O., El-Sayed, N., Srikantiah, P., Luby, S.P., Earhart, K., Mahoney, F.J., 2007. Brucellosis as a cause of acute febrile illness in Egypt. Trans. R. Soc. Trop. Med. Hyg. 101, 707–713. Mainar-Jaime, R.C., Litgh-Pereira, P.L., Epp, T., Waldner, C., 2005. The application of spatial analysis tools in small-ruminant brucellosis eradication programs in Northern Spain. Intern. J. Appl. Res. Vet. Med. 3, 179–188. Minas, A., 2006. Control and eradication of brucellosis in small ruminants. Small Rumin. Res. 62, 101–107. Moreno, E., Cloeckaert, A., Moriyón, I., 2002. Brucella evolution and taxonomy. Vet. Microbiol. 90, 209–227. Ministry of Agriculture and Land Reclamation, 2005. Agricultural census in Egypt, Economic Affairs Sector. Muma, J.B., Samui, K.L., Oloya, J., Munyeme, M., Skjerve, E., 2007. Risk factors for brucellosis in indigenous cattle reared in livestock–wildlife interface areas of Zambia. Prev. Vet. Med. 80, 306–317. Newcombe, R.G., 1998. Two sided confidence intervals for the single proportion: comparison of seven methods. Statist. Med. 17, 857–872. Pappas, G., Papadimitriou, P., Akritidis, N., Christou, L., Tsianos, E.V., 2006. The new global map of human brucellosis. Lancet Infect. Dis. 6, 91–99. Radostits, O.M., Arundel, J.H., Gay, C.C., Blood, D.C., Hinchcliff, K.W., 2000. Veterinary medicine: a textbook of the diseases of cattle, sheep, pigs, goats and horses. WB Saunders Company, Philadelphia. Ramirez-Pfeiffer, C., Nielsen, K., Smith, P., Marin-Ricalde, F., RodriguezPadilla, C., Gomez-Flores, R., 2007. Application of the fluorescence polarization assay for detection of caprine antibodies to Brucella melitensis in areas of high prevalence and widespread vaccination. Clin. Vaccine Immunol. 14, 299–303. Refai, M., 2002. Incidence and control of brucellosis in the Near East region. Vet. Microbiol. 90, 81–110. Rivera, S.A., Ram rez, M.C., Lopetegui, I.P., 2002. Eradication of bovine brucellosis in the 10th Region de Los Lagos, Chile. Vet. Microbiol. 90, 45–53. Robinson, A., 2003. Guidelines for coordinated human and animal brucellosis surveillance. FAO, Rome. Samaha, H., Al-Rowaily, M., Khoudair, R.M., Ashour, H.M., 2008. Multicenter study of brucellosis in Egypt. Emerg. Infect. Dis. 14, 1916–1918. WHO, 2009. Seven Neglected Endemic Zoonoses – some basic facts. http://www.who.int/zoonoses/neglected zoonotic diseases/en/ (last accessed May 2009). OIE, 2004. Manuals of Diagnostic Tests and Vaccines for Terrestrial Animals, Chapter 2.3.1 (Caprine and Ovine brucellosis). http://www.oie.int/fr/normes/mmanual/A 00069.htm (last accessed 07.01.2010).
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Ruminant brucellosis in Upper Egypt (2005–2008) Y.M. Hegazy a,∗ , B. Molina-Flores b,d , H. Shafik c , A.L. Ridler e , F.J. Guitian f a
Department of Animal Medicine, Veterinary College, Kafrelsheikh University, Kafr Elsheikh, Egypt Egyptian Spanish Project for the Control of Brucellosis in the Upper Egypt, Spanish Agency for International Cooperation and Development, 21 El-Mahad El Swissry, Cairo, Egypt c General Organization for Veterinary Services, Ministry of Agriculture and Land Reclamation, Nadi El Said, Dokki 12311, Cairo, Egypt d Regional Animal Health Center for North Africa, Food and Agriculture Organization of the United Nations. 43, Avenue Kheireddine Pacha – 1002 Tunis, Tunisia e Institute of Veterinary, Animal & Biomedical Sciences IVABS, Tennent Drive, Massey University, 4442 New Zealand f Veterinary Epidemiology and Public Health Group, Department of Veterinary Clinical Sciences, The Royal Veterinary College, University of London, North Mymms, Hertfordshire AL9 7TA, UK b
a r t i c l e
i n f o
Article history: Received 25 March 2010 Received in revised form 9 April 2011 Accepted 11 May 2011 Keywords: Brucellosis Control program Households Ruminants Spatial analysis Upper Egypt
a b s t r a c t Brucellosis is endemic among humans and ruminant in Egypt and recent reports suggest that its incidence may be increasing. In this study we describe the frequency of brucellosis among different ruminant species in Upper Egypt and its spatial distribution using the data generated by a large-scale control campaign undertaken between 2005 and 2008. A total of 120,090 individual animals of different ruminant species were tested during the campaign. The true proportions of brucellosis were estimated as 0.79% (CI: 0.71%–0.87%), 0.13% (CI: 0.08%–0.18%), 1.16% (1.05%–1.27%) and 0.44% (0.34%–0.54%) among cattle, buffaloes, sheep and goats respectively. We estimated that 0.2% (CI: 0.16%–0.23%) of households in the study area keep at least one seropositive animal. Spatial autocorrelation of the proportions of seropositive households and seropositive animals was assessed using Global Univariate Moran’s I and Local Univariate LISA. These analyses showed that the distribution of seropositive animals has considerable spatial heterogeneity with clustering in the northern governorates of the study area. Our results show that brucellosis is widespread and heterogeneously distributed in Upper Egypt. At the current level of available resources it is very unlikely that test and slaughter could be implemented with the intensity needed to be effective and other control measures that could replace or complement the test and slaughter policy in place should be considered. Also, this study illustrates some of the challenges faced by bilateral projects that have to accommodate an externally funded intervention with an ongoing national official disease control program. © 2011 Elsevier B.V. All rights reserved.
1. Introduction Brucellosis is one of the world’s most widespread zoonotic diseases caused by species of the genus Brucella (WHO, 2009; Moreno et al., 2002). Infection in ruminants causes economic losses due to abortion, infertility and pre-
∗ Corresponding author. Tel.: +20 2402790113. E-mail address: [email protected] (Y.M. Hegazy). 0167-5877/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.prevetmed.2011.05.007
mature culling (Radostits et al., 2000). Control of brucellosis depends generally on test and isolation/slaughter of positive animals, vaccination of susceptible animals and control of animal movements (Corbel, 1997). Various countries all over the world have implemented different control policies using combinations of these control measures, with reported success in some countries and failure in others (Cutler et al., 2005; Blasco, 1997). In Egypt, since 1981, the General Organization of Veterinary Services (GOVS) has run a control program which
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is currently based on testing female ruminants older than 6 months of age with slaughter of serologically positives, and voluntary vaccination of calves using Brucella abortus S19 vaccine and Brucella melitensis Rev 1 vaccine for lambs and kids (Refai, 2002). Despite almost 30 years of implementation of the control program, brucellosis remains endemic among ruminants and humans in Egypt and recent reports suggest that the incidence of human infection may be increasing (Jennings et al., 2007). Given that infected animals are the source of human infection, the increasing incidence of human brucellosis probably reflects a similar trend in domestic animals. Unfortunately, reliable data on the frequency and distribution of ruminant brucellosis in Egypt is lacking. Perhaps a reason why the official control program has limited success and has not been always fully adhered to is that the strategies were not adequate for the baseline frequency of disease (Hegazy et al., 2009). Indeed, it is generally accepted that depending on the prevalence/incidence of brucellosis among different animal species, different combinations of control strategies may be appropriate (Robinson, 2003). A better understanding of the frequency of infection and its distribution across geographic areas may help tailor the control program to the situation and may contribute to both its actual implementation as per official guidelines and its impact in reducing the frequency of infection in livestock and humans. This is the overall aim of the present study, which we have tried to achieve for the Upper Egypt region, an area in the valley of the Nile river, south of the delta, which contains around 32% and 39% of the total Egyptian large and small ruminant population respectively (Ministry of Agriculture and Land Reclamation, 2005). In this area it is estimated that more than 90% of the large ruminants are kept by households owning between 1 and 5 cattle and/or buffaloes which are often in close contact with sheep and goats (Aidaros, 2005). Accurate and unbiased estimates of the frequency of brucellosis in the region are lacking, but a recent study in two of the Upper Egypt governorates found between 4.5 and 7% of seropositive animals among different ruminant species (Samaha et al., 2008). Lack of information on the target population and sampling procedure for villages/farms and animals within them make the results of this study difficult to extrapolate to the general population. However, if the results are not a gross overrepresentation of the individual-level true prevalence of infection, they seriously question the appropriateness of the current control strategy based on test and slaughter of positive animals and voluntary vaccination of young animals. Only if the prevalence was considerably lower and enough resources were available for frequent testing of a large enough proportion of the population would such a program be likely to succeed (Hegazy et al., 2009). In the current situation, estimates of the frequency of infection are urgently required to make a preliminary assessment of the appropriateness of the control strategy in place. In 2005, a 3-years long Egyptian-Spanish bilateral project for the control of brucellosis in Upper Egypt was initiated and jointly implemented by the GOVS and the Spanish Agency for International Cooperation and
Development (AECID). The project aimed at reducing the incidence of brucellosis in the ruminant population of Upper Egypt by providing support to the GOVS control program. Specifically, the proportion of adult ruminants tested in each governorate would be increased concurrently with a vaccination campaign. Assuming probabilistic selection of animals to be tested and the slaughter of those found positive it was expected that the enhanced control program would (i) generate better prevalence estimates (ii) reduce the incidence of ruminant brucellosis in the region. The primary objective of this study is to describe the frequency of brucellosis among different ruminant species in Upper Egypt and its spatial pattern using the results of the 2005–2008 brucellosis control campaign carried out in this area. A secondary objective is to describe this bilateral control program and in particular the main challenges and lessons learned from the implementation of a large scale brucellosis control program in an endemic area. 2. Materials and methods 2.1. Study area and control campaign activities The study area consisted of the 7 Upper Egypt governorates of Beni Sueif, Al Minia, Assuit, Sohag, Quena, Luxor and Aswan. The governorates of Al Fayoum and Al Giza as well as the southern area of Cairo are often included as part of Upper Egypt but for purpose of this study they are excluded. Within these 7 governorates only 6.4% of the land area is populated, the remainder is desert (Fig. 1). Our study was restricted to the populated area of the 7 governorates, which includes 55 administrative units (districts) with 430 veterinary units. Veterinary units are governmental units distributed all over Egypt (around 1500 as mentioned by Aidaros, 2005) managed by one or more official veterinarians. These units are responsible for supplying different veterinary services to the livestock sector including the application of mandatory interventions. The study area represents 13% of the total populated area of Egypt. The control campaign was initiated in October 2005 and finished in October 2008. It was planned for this campaign to consist of 2 main activities: - First, yearly serological testing against Brucella spp. of a randomly selected 10% of the total adult ruminant population in each governorate (130,999 total animals to be tested from the whole of Upper Egypt, yearly). The number of animals to be tested every year was calculated in order to allow yearly estimation of the prevalence of seropositive animals of the different species (cattle, buffalo, sheep, goats) in each governorate, with 90% confidence and 15% precision, using as expected prevalence values the % of seropositive animals among those tested as a result of the GOVS control activities in 2003, previous to the start of the project. The resulting number of animals of the different species to be sampled in each governorate was divided equally among the local veterinary units of the governorate. The veterinarian in charge of the veterinary unit was asked to select randomly the allocated number of samples and to collect the samples from all villages served by the clinic and from differ-
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2.2. Data sources The available data for this study consisted of information on the sex, age, breed, farm/household location (governorate, district and village), owner identification, date of sampling and results of the serological tests for all individual animals sampled as part of the control campaign between October 2005 and January 2008. This information was stored in a relational database that was developed as part of the campaign. Each veterinary unit was provided with a computer supplied with software specifically built by the project for recording of the required data for each animal. The veterinarians received training on how to collect the field data using predesigned forms and how to use the database and introduce data remotely. 2.3. Data analysis For those animals that were tested more than once (4113 animals), only the first record was used in the analysis. 2.4. Descriptive statistics Descriptive statistics were obtained for the key variables in relation to the different aspects of the campaign such as numbers of tested animals, villages and local veterinary units involved in the campaign. Fig. 1. Map of Upper Egypt showing the location of the seven governorates of the study area and their populated and desert areas.
ent farms within the village, however, random sampling was not fully adhered to. This was attributed to lack of incentives for veterinarians and poor cooperation from farmers. Also, no further explanation was provided with regard to what random selection involves and how it can be performed across different levels (selection of households or farms within villages and of individual animals within households or farms). This initial annual target was revised soon after the start of the campaign to the same number of animals to be tested but across all the 3 years of the campaign rather than annually. Such testing would be carried out while the GOVS would continue to be responsible for slaughtering of positive reactors and compensating the owners. Serological testing consisted of Rose Bengal Plate test (RBPT) followed by Complement Fixation test (CFT) for animals seropositive to the former test (series interpretation), as officially approved by the national veterinary services in Egypt in accordance with the OIE manual of standard diagnostic tests and vaccines (World Organization for Animal Health, 2004). - Second, mass vaccination of all female cattle and buffaloes with Brucella abortus S19 at a dose of 1010 and of all sheep and goats (males and females) with Brucella melitensis Rev1 at a dose of 109 .
These activities were to be undertaken by 964 veterinarians and 2396 assistants from 430 local veterinary units.
2.5. Seroprevalence of brucellosis at individual animal level The apparent prevalence was calculated as the number of animals found to be seropositive divided by the total number of animals sampled during the study period. It was obtained both, for all species combined and by species. The true proportion of seropositive animals among those tested (TP) and the true proportion of seropositive animals by species (STP) were calculated after adjusting for the combined sensitivity (CSe) and combined specificity (CSp) of the serological tests from the apparent prevalence (AP) as TP = (AP + CSp − 1)/(CSe + CSp − 1). The values of CSe and CSp of the testing strategy were obtained, assuming series interpretation, as the most likely values from triangular distributions for CSe = SeRBT × SeCF and for CSp = 1 − ((1 − SpRBT ) × (1 − SpCF )); where SeRBT , SpRBT, SeCF, SpCF are the sensitivity and specificity of RBPT and CFT respectively. A very wide range of values for Se and Sp are presented in the published literature; to account for this variability, the values of Se and Sp of individual tests were obtained by random selection from uniform distributions with minimum and maximum values derived from the literature: 0.72 < SeRBT < 1; 0.8 < SpRBT < 1; 0.81 < SeCF < 1; 0.80 < SpCF < 1 (Blasco et al., 1994; Gall and Nielsen, 2004; Ramirez-Pfeiffer et al., 2007). Ten-thousand values of the sensitivity and specificity of both serological tests were chosen from the uniform distributions to generate the distributions of CSe and CSp. Values of CSe and CSp were assumed to be the same across species. 95% confidence intervals (CI) around the estimated proportion were obtained as a measure of precision of the estimates
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(Newcombe, 1998). The STP was also calculated by year. The true proportion of seropositive animals by district (DTP) was estimated the same way as TP and STP. 2.6. Seroprevalence of brucellosis at household level For each district, the proportion of “tested households” with serologically positive animals (household-level district apparent prevalence: HDAP) was obtained. Any household for which one or more tested animals were found seropositive was considered a seropositive household. Information on herd/flock size for those households in which animals were sampled was not available but the vast majority of households keeping ruminants in this part of Egypt have just a few animals (Aidaros, 2005). It was assumed in this study that the total number of animals tested from a single owner was the total size of the household herd/flock – i.e. it was assumed that once a household was included in the campaign all its animals had been sampled. Using estimated values of householdlevel sensitivity and specificity (details below) the true proportion of seropositive households by district (HDTP) was estimated from the apparent proportion (HDAP) as HDTP = (HDAP + HCSp − 1)/(HCSe + HCSp − 1), where HCSp is the household-level combined specificity and HCSe the household level combined sensitivity. The apparent and true proportion of households with seropositive animals for brucellosis in each governorate of Upper Egypt (HGAP and HGTP) were calculated in the same way as HDTP. The apparent and the true overall prevalence (HAP and HTP) for the whole area of Upper Egypt were calculated in the same way. 2.7. Simulation of the likely performance of the testing strategy at household level The likely ability of the testing strategy to correctly identify households with and without infected animals was assessed by means of simulation. For purpose of this simulation it was assumed:
of infected households with seropositive animals, noninfected households with seropositive animals, infected households without seropositive animals and non-infected households without seropositive animals were used to calculate the HCSe and HCSp. 2.8. Spatial distribution of seropositive animals and households To visualize the geographic distribution of ruminant brucellosis in Upper Egypt, two choropleth maps representing the estimated true prevalence of seropositive animals and the estimated true prevalence of seropositive households per district were created using Arc GIS 9.2 (ESRI 2006). To assess the extent to which the status of a district with regard to brucellosis (as defined by the proportion of seropositive animals and the proportion of its households with seropositive animals) was spatially correlated within the study area, we calculated the global univariate Moran’s I considering any district that shares any common point with another district as neighbor (first order queen contiguity). Since some of the districts had few households and animals tested, numbers of animals and households sampled per district were used in an empirical Bayes smoothing method to adjust for underlying population structure (Henning et al., 2009). Inference for the significance of the value of Moran’s I obtained was assessed by running this analysis for 999 permutations. In order to identify areas with either homogeneously high or low seroprevalence of brucellosis we used Local Univariate LISA (Local Indicators of Spatial Association) to examine the local level of spatial autocorrelation. The same settings as for the Global Univariate Moran’s I were used; significant clusters were identified after running the analysis for 999 permutations at P < 0.05. These analyses were carried out using GeoDa 0.9.5-i.5 (http://geoda.uiuc.edu/). 3. Results 3.1. Proportion of seropositive animals
- All individual animals in the population were equally likely to be infected regardless of their aggregation within households (intracluster correlation coefficient () = 0). - All households were equally likely to be sampled with probability = 0.1. - The distribution of number of animals per household differed between districts. Household size was assumed to follow a triangular distribution with parameters varying between districts; minimum and maximum values where derived from the distribution of animals sampled from the same owner by district. When a household with n animals is sampled, any number of animals from 1 to n can be sampled with equal probability. - The combined sensitivity and specificity of the tests interpreted in series at individual animal level were used as calculated above. The simulation was carried out in @Risk software (version 3.5d, Palisade Corporation, Newfield, NY, USA). The simulation was run for 10,000 iterations. The total number
A total of 120,077 individual animals of different ruminant species were tested between October 2005 and January 2008 and had complete information. Twelve percent of the local veterinary units did not participate in animal sampling during this period, the remaining 88% veterinary units collected samples from 54% of the villages in the study area, which suggests some clustering of sampling by village (Table 1). The CSe and CSp of the series combination of the two tests ranged triangularly from 64% to 92% and 97% to 100% respectively with most likely values found to be 78% and 99–100% respectively. A total of 1071 animals were found seropositive to RBPT and 693 of them were found positive to CFT. The TP was estimated as 0.74% (CI: 0.69%–0.79%). By species, the STP were estimated as 0.79% (CI: 0.71%–0.87%), 0.13% (CI: 0.08%–0.18%), 1.16% (1.05%–1.27%) and 0.44% (0.34%–0.54%) for cattle, buffaloes, sheep and goats respectively. Only data from years 2006 and 2007 will be used in the analyses by year (the results of 2005 and 2008 were
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the proportion ranging from 0.1% to 3.0% in the different governorates and years of the campaign (Table 2). Based on our simulations, the HCSe and HCSp, ranged from 47.0% to 73.0% and from 95.0% to 97.0% respectively according to district. If our assumptions hold true, the overall HTP would be 0.2% (CI: 0.16%–0.23%). The HDTP were calculated and showed a large fluctuation (from 0.1% to 7.7%) between districts as shown in Fig. 3. The HGAP and HGTP for different governorates are shown in Table 2.
3.3. Spatial distribution of seropositive animals and households Fig. 2. Estimated true proportions of serologically positive individuals of different ruminant species against Brucella spp. in Upper Egypt 2006–2007.
omitted, as there were low numbers of animals and households sampled in this period; 21 animals in 2005 and 1859 animals in 2008). In 2006 the numbers of tested animals were 30,154 cattle, 6615 buffaloes, 19,046 sheep and 7252 goats, while for 2007, 19,004 cattle, 11,438 buffaloes, 13,987 sheep and 10,701 goats. The STP in 2006 and 2007 by species are presented in Fig. 2 and the overall DTP for the combined result of years 2006 and 2007 is shown in Fig. 3. 3.2. Seroprevalence at household level Overall, 1.2% (CI: 1.1%–1.3%) of the total households tested had seropositive animals against Brucella spp., with
Districts in the northern governorates of the study area (Beni sueif and Al Minia) had the highest proportions of positive animals. There was a tendency towards a declining proportion of seropositive animals from Northern to Southern districts in the study area (Fig. 3). The value of the Global Moran’s I using empirical Bayes smoothing for individual animal prevalence was 0.58 (P < 0.001) indicating that there was a significant spatial autocorrelation (clustering) of the proportion of positive individual animals between districts. The results of location of clusters identification using LISA with spatial empirical Bayes indicated that there was significant spatial clustering (P < 0.05) between districts with high proportions of seropositive animals in Beni Sueif and Al Minia governorates (northern part of the study area). On the other hand, there was a tendency towards increasing proportion of seropositive households from Northern to Southern districts in the study area (Fig. 3). The
Fig. 3. Distribution of estimated true proportion of individual ruminants (A) and households (B) seropositive against Brucella spp. among Upper Egypt districts 2006–2007.
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Fig. 4. LISA cluster maps for identification of districts with similar proportions of (A) ruminants seropositive against Brucella spp. and (B) households seropositive against Brucella spp. in Upper Egypt, 2006–2007.
value of the Global Moran’s I using empirical Bayes smoothing for household prevalence was – 0.013 (P < 0.445) which indicates a non significant clustering of seropositive households. Results of local cluster identification showed that there was clustering of districts with low proportions of seropositive households in Beni Sueif governorate (Fig. 4). 4. Discussion Brucellosis is an endemic disease of humans and ruminants throughout most of the Middle East and some countries of the Mediterranean basin (Refai, 2002; Benkirane, 2006; Pappas et al., 2006). Its economic and public health importance has lead countries in the area to establish different control programs (Refai, 2002). In Egypt, the GOVS has run a program for the surveillance and control of ruminant brucellosis since 1980. The main elements of this control program consist of serological testing every 6 months of all female cows, buffaloes, sheep and goats over 6 months old and valuable bulls, rams and bucks using RBPT and CFT with slaughtering of seropositive animals and compensation of the owners. This is accompanied by voluntary vaccination of calves, lambs and kids which are serologically negative at 3–6 months age, using S19 vaccine for cattle and buffaloes and Rev1 vaccine for sheep and goats (Refai, 2002). A recent study showed that no more than 7% of target animals were tested every year (Hegazy et al., 2009), because of lack of farmers and veterinarians incentives and insufficient funding to sustain the application of this policy. There are no accurate figures for the actual coverage of the official vaccination program, but from dis-
cussions with the project manager of the campaign and with veterinarians from the Egyptian GOVS and on the basis of figures previously reported to the OIE, the voluntary vaccination program appears to have very limited coverage and unlikely to result in the vaccination of more than 1% of the calves, lambs and kids. These factors have resulted in the inability of this program to decrease the prevalence of brucellosis in Egypt (Refai, 2002; Hegazy et al., 2009, 2011). It has also resulted in the absence of reliable estimates of the frequency of infection and how it varies across species, production systems and geographic areas. The main reason why such estimates are not available is lack of a formal sampling strategy; it has been suggested that, at least for some parts of the country, the implementation of the test and slaughter campaign relies on haphazard sampling concentrated in areas that are accessible and with huge variations in its intensity across species year to year (Hegazy et al., 2009). One of the objectives of the 2005–2008 control campaign was to decrease the prevalence of brucellosis among different ruminant species by means of obligatory whole herd vaccination. The other objective was to generate data that could be used to estimate the seroprevalence of brucellosis in Upper Egypt and, accordingly, the campaign aimed at probabilistic sampling of individual animals. In this study, we attempted to fulfill some of the knowledge gaps with regard to the frequency and distribution of brucellosis in Upper Egypt by analyzing the large amount of data generated by the 2005–2008 control campaign. Over 3 years (October 2005 to January 2008), the campaign succeeded in testing around 9% of the total adult
Table 1 Total numbers of local veterinary units in Upper Egypt and distribution of these veterinary units regarding the proportions of their served villages which are sampled for brucellosis as part of the Egyptian-Spanish bilateral project for the control of brucellosis in Upper Egypt in 2006 and 2007. Total number of Veterinary Units
Number of Veterinary Units which did not test
*
Veterinary Units which tested only one village
Veterinary Units which tested <50% of their associated villages
Veterinary Units which tested <75% of their associated villages
Total number of villages
Number of villages sampled
Beni Sueif Al Minia Assuit Sohag Quena Luxor Aswan Total
76 90 77 80 48 11 48 430
2 (3%) 3 (3%) 25 (32%) 14 (17%) 1 (2%) 0 (0%) 5 (10%) 50 (12%)
16 (21%) 8 (9%) 15 (19%) 16 (20%) 2 (4%) 0 (0%) 2 (4%) 59 (14%)
32 (42%) 26 (29%) 44 (57%) 42 (53%) 7 (15%) 1 (9%) 10 (21%) 162 (38%)
52 (68%) 54 (60%) 58 (75%) 64 (80%) 20 (42%) 3 (27%) 20 (42%) 271 (63%)
466 612 359 570 334 96 252 2689
227 (49%) 358 (58%) 138 (38%) 253 (44%) 243 (73%) 70 (73%) 158 (63%) 1447 (54%)
*
These Veterinary units serve more than one village.
Table 2 Results of serological testing against Brucella spp. of households containing ruminants as a part of the Egyptian-Spanish bilateral project for the control of ruminant brucellosis in Upper Egypt in 2006 and 2007. Year
2006
Governorate Total number of households tested Beni Sueif Al Minia Assuit Sohag Quena Luxor Aswan Total
6703 3868 2778 2658 4236 478 1748 22,469
2007 Number of negative households 6502 3822 2764 2651 4224 477 1747 22,187
Number and % of positive households 201 (3.0%) 46 (1.2%) 14 (0.5%) 7 (0.3%) 12 (0.3%) 1 (0.2%) 1 (0.1%) 282 (1.3%)
Total number of households tested 4428 3442 280 3578 3494 1684 909 17,822
Number of negative households 4294 3412 280 3585 3486 908 906 17,638
Number and % of positive households 134 (3.0%) 30 (0.9%) 0 (0.0%) 7 (0.2%) 8 (0.2%) 2 (0.1%) 3 (0.3%) 184 (1.0%)
Total number of tested households 11131 7310 3058 6243 7730 2162 2657 40,291
Apparent proportion of True proportion of positive households positive households %(HGAP) %(HGTP) (CI) 3.0% (2.7–3.3) 1.0% (0.8 –1.2) 0.5% (0.3–0.7) 0.2% (0.1–0.3) 0.3% (0.2–0.4) 0.1% (0.0–0.3) 0.2% (0.0–0.4) 1.2% (1.1–1.3)
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Governorate
0.3% (0.2–0.4) 0.1% (0.0–0.2) 0.4% (0.2–0.6) 0.2% (0.1–0.3) 0.1% (0.0–0.2) 0.1% (0.0–0.2) 0.2% (0.0–0.4) 0.2% (0.16–0.23)
CI, confidence interval.
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ruminant population in Upper Egypt with slaughtering of 125 seropositive animals. The remaining seropositive animals were not slaughtered because of the lack of available funding to compensate their owners. The estimated true proportions of seropositive animals in the study area ranged from 0.13% in buffaloes to 1.16% in sheep. This seroprevalence estimate should not have been substantially affected by the vaccination campaign, given the low coverage. However, caution should be taken when interpreting these estimates which may not reflect the actual proportion of positive animals in the study area because of some weaknesses: despite the instructions given to the local veterinarians that random selection should have been used, they were not given further explanations on what random selection actually implies and how it can be achieved. Furthermore, veterinarians were expected to carry out the sampling as part of their normal duties without additional incentives and no supervision or formal control of the way they conducted the sampling was in place. These limitations raise considerable doubts as to the extent to which probabilistic selection was achieved, limiting our ability to extrapolate the obtained results to the reference livestock population of Upper Egypt. We estimated that 0.2% of all households in the study area had seropositive animals. This figure accounts for the inadequate performance of diagnostic tests (based on our assumptions) but, similarly to the animal-level estimates, it may be affected by selection bias due to non-random selection of households and animals within households. However, we believe that household-level estimates are probably less influenced by departures from probabilistic sampling and aggregation of samples within certain clusters than individual animal estimates. Overestimation of the true proportion of positive households due to selection bias cannot be ruled out, but we consider this to be unlikely. Based on our experience in the area, selection bias would be due to overrepresentation of households nearby the veterinary clinics and therefore unrelated to brucellosis status. The main Brucella species responsible for infection in ruminants are Brucella abortus and Brucella melitensis. In this research, we did not identify the Brucella species in infected animals. However, the main isolate in all animal species and humans in Egypt is Brucella melitensis (Refai, 2002) and there is evidence that suggests that cattle act as spill-over hosts of this species (Hegazy et al., 2011). Our results suggest that ruminant brucellosis is not homogeneously distributed across the study area, with higher prevalence of infection in northern districts. A possible explanation may be the location of these districts closer to the large animal markets in Cairo where animals from different sources, including the Nile delta mix and are redistributed (Muma et al., 2007). A second factor that could contribute to the higher prevalence of infection observed in the northern governorates of Upper Egypt is that this area has one of the highest densities of ruminants in Egypt, in particular sheep and goats, which are often reared in movable flocks (Ministry of Agriculture and Land Reclamation, 2005). These types of flocks are particularly common in Al Minia governorate (Fayed, personal communication).
Prevalence estimates are the cornerstone for deciding the appropriate control policy for brucellosis and for testing its efficiency. Despite the vast amount of data it generated, the 2005–2008 campaign does not provide prevalence estimates that could assertively be regarded as unbiased. At the same time, the results obtained provide some insight on the likely disease distribution among different ruminant species and different geographic areas in Upper Egypt. As for the efficiency of the campaign in reducing the frequency of infection in the target population, only a small proportion of the ruminant population was tested every year. Testing such a small proportion of the population with slaughtering of positive animals would allow the infection to persist. Consequently, the cost-effectiveness of the campaign seems to be, at least, questionable. Furthermore, lack of funding for compensation of livestock owners did not allow for all animals found positive to be culled. The study illustrates some of the challenges resulting from the need to accommodate the activities of an externally funded 3-year intervention and an ongoing official disease control program. Soon after the project was started its objectives were revised. The original intention of achieving probabilistic sampling of 10% of the total ruminant population of Upper Egypt every year was soon replaced for the same proportion but during the whole 3-year duration of the program and the vaccination campaign had to be delayed until the very end of the project because of logistic difficulties (it is currently being undertaken). A major challenge of this particular control program has been that a sampling strategy aimed at gaining knowledge on the frequency of infection (as suggested by the intention of randomly selecting animals to be tested stated in the protocol of the campaign) had to be coordinated with the official test and slaughter control policy. Some of the insights into the situation of ruminant brucellosis in Upper Egypt gained from this study should be taken into account when designing the next phases of the official control campaign. The current test and slaughter policy is a reasonable strategy for the control of brucellosis in Upper Egypt only if our estimates for the proportion of seropositive animals and households are not heavily biased downwards and if a sufficient proportion of the ruminant population could be tested frequently enough (Corbel, 2006; Hegazy et al., 2009) accompanied with slaughtering of seropositive animals. Even with external assistance, it has proved challenging to test 10% of the ruminant population in a 3-year period, therefore it seems unlikely that with the current level of resources test and slaughter could be implemented with the intensity needed to significantly reduce the prevalence of infection among ruminants in the area. For that reason, the national control program should also consider other control strategies that can replace or complement test and slaughter such as compulsory vaccination and more strict application of quarantine and movement control (Minas, 2006). The observed spatial heterogeneity may offer opportunities for targeted interventions in the ruminant population. Interventions that could minimize the spread of the infection from the Northern governorates of Upper Egypt to non-infected areas or areas with lower prevalence may be a cost-effective strategy (Mainar-Jaime et al., 2005). Geographic zoning as a
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policy for brucellosis control and/or eradication has been successfully implemented in some countries such as Chile, where brucellosis eradication programs were initiated in regions with low cattle density and low prevalence, and then extended gradually to other regions (Rivera et al., 2002). Similar strategies could be particularly appropriate in countries with scarce economic resources such as Egypt. A possible alternative to the current control program could be compulsory vaccination in geographic areas with high seroprevalence (FAO/WHO/OIE, 1993). For such interventions to succeed in reducing the incidence of ruminant brucellosis additional measures such as control of animal movement would be required. Crucially, as the 2005–2008 campaign clearly shows, the success of any control strategy for brucellosis in Upper Egypt has to consider the perceptions and incentives of farmers and veterinarians and try to engage them. The spatial aggregation of districts with relatively high proportions of seropositive animals in the Northern governorates of Upper Egypt, suggests that the risk of human exposure to Brucella spp. maybe particularly high among rural households in this region. Public health services should probably pay particular attention to the rural population of this area during their campaigns for prevention of human brucellosis. 5. Conclusion Brucellosis is widespread among different ruminant species in Upper Egypt, where we estimate that 0.2% of households keep seropositive ruminants. Infection is heterogeneously distributed, with highest prevalence in the ruminant population of the Northern areas and as a result higher risk of human exposure. At the current level of available resources it is very unlikely that test and slaughter could be implemented with the intensity needed to be effective and other control measures should be considered. Acknowledgements We would like to acknowledge the staff members of the General Organization of Veterinary Services in Cairo and ˜ de Cooperación Internacional para el the Agencia Espanola Desarrollo for their help with the collection of data and the permission to use these data in this study. References Aidaros, H., 2005. Global perspectives – the Middle East: Egypt. Rev. Sci. Tech. Off. Int. Epiz. 24, 589–596. Benkirane, A., 2006. Ovine and caprine brucellosis: world distribution and control/eradication strategies in West Asia/North Africa region. Small Rum. Res. 62, 19–25. Blasco, J.M., 1997. A review of the use of B. melitensis Rev 1 vaccine in adult sheep and goats. Prev. Vet. Med. 31, 275–283.
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