This section summarizes Identification, Serological and other tests, Vaccines and Diagnostic Biologicals, based on Chapter 2.4.2 of the OIE Manual of Standards for Diagnostic Tests and Vaccines (2000).
Brucella melitensis (biovars 1, 2 or 3) is the main causative agent of caprine and ovine brucellosis. Sporadic cases caused by B. abortus have been observed, but clinical disease is uncommon. B. melitensis occurs in the Mediterranean region, but infection is also widespread, especially where small ruminants are the major livestock species. Canada and the USA are believed to be free, as are northern Europe, Southeast Asia, Australia and New Zealand.
Clinically, the disease is characterized by one or more of the following signs: abortion, retained placenta, orchitis, epididymitis and, rarely, arthritis.
B. melitensis is highly pathogenic for humans, causing one of the most serious zoonoses in the world. All infected tissues, cultures and potentially contaminated materials should be handled under conditions for biohazard containment.
Presumptive evidence of Brucella is provided by the demonstration, by modified acid-fast staining of organisms, of Brucella morphology in abortion material or vaginal discharge, especially if supported by serological tests. The recently developed polymerase chain reaction methods provide additional means of detection. Whenever possible, Brucella sp. should be isolated by culture - using selective media - from uterine discharges, aborted foetuses, udder secretions or selected issues, such as lymph nodes, testes or epididymides. Species and biovars should be identified by phage lysis, and by cultural, biochemical and serological criteria.
The Rose Bengal plate agglutination, complement fixation and indirect ELISA tests are usually recommended for screening flocks and individual animals. The complement fixation test is the only test prescribed for confirmation and international trade, but other tests, such as the immunodiffusion and competitive ELISA, are useful for confirmation purposes. The serum agglutination test (SAT) is not considered reliable for use in small ruminants. For pooled samples, there are no useful tests equivalent to the milk ring test in cattle. The brucellin allergic skin test can be used as a screening or complementary test in unvaccinated flocks, provided that a lipopolysaccharide-free and standardized antigen preparation is used. Results must then be interpreted in relation to the clinical signs, history, and the results of serological or cultural examination.
B. melitensis strain Rev.1 live vaccine is recommended to immunize sheep and goats at risk of infection from Brucella. Production of Brucella antigens of Rev.1 vaccine is based on a seed-lot system. Seed cultures to be used for antigens for serological and allergic skin tests and for vaccines should originate from reference centres. They must conform to minimum standards for viability, smoothness, residual infectivity and immunogenicity, if applicable. Antigens for serological tests should be standardized against reference sera calibrated against the International Standard Anti-Brucella abortus Serum, and the procedures and interpretations should conform to international recommendations. Allergen preparations should be free of lipopolysaccharide to prevent anti-lipopolysaccharide antibody production and local inflammatory reactions.
During this phase, the magnitude and distribution of the problem should be determined (as discussed earlier).
Voluntary investigation of abortion incidents and orchitis/epididymitis lesions, and submission to a diagnostic laboratory for culture (passive).
Off farm surveillance
Percentage of abortion incidents confirmed as brucellosis (passive).
Bacteriological and serological examination of tissues and blood from breeding age sheep and goats at slaughter (active).
Note that in some countries, home or illegal slaughter may constitute a high percentage of all animals killed so that legal-slaughter-based samples may not always be representative.
Where brucellosis is present at high rates, especially in developing countries, sheep and goats are usually managed under extensive transhumant or nomadic systems. Under these conditions, B. melitensis cannot be eradicated by test and slaughter alone, and a vaccination programme has to be applied to reduce the risk of spread of the disease. Live Rev.1 B. melitensis vaccine is considered to be the best available for use in small ruminants. Originally it was believed that exclusive vaccination of the young replacement animals annually for 5-6 years (the usual productive life-span of these species) would be sufficient, based on the assumption that the Rev.1 vaccine resulted in lifelong immunity. However, this strategy has failed in both developed and developing countries. Possible explanations for these failures included (i) a low vaccination coverage because owners keep introducing replacements continuously throughout the year, (ii) vaccine quality; or (iii) decreased immunity with age. Accordingly, whole flock or herd vaccination every two years is an alternative to control B. melitensis infection under extensive management conditions. The major risk with the use of Rev.1 at standard doses (1-2 ´ 109 CFU/dose) administered subcutaneously to adult sheep and goats is that some pregnant animals may abort. While parturition is seasonal, a few animals may be pregnant at any one time. Reduction of the dose to 104-107 CFU/dose given subcutaneously has been used in an effort to reduce the risk of abortion, but overall the results of experimental and field studies have shown that the resulting immunity is inadequate. Conjunctival vaccination with the standard dose of Rev.1 is safer than subcutaneous vaccination, but is not safe enough to be applied regardless of the pregnancy status of the animals, and should be used at a time of the year when the majority of the sheep are at least risk, i.e. during late lambing or kidding season, or during lactation.
Test random samples of animals and herds 2-3 weeks after vaccination using the Rose Bengal test to evaluate vaccine response and coverage. Also check identification (active).
Monitor abortion incidents as in the previous phase, or carry out active surveillance on selected or sentinel flocks and herds to determine abortion rates. Confirm using bacteriology whether abortion is due to B. melitensis. As the vaccine strain (Rev.1) may cause abortion also, all isolates should be typed especially to differentiate Rev.1 from field strains of B. melitensis biovar 1.
Monitor randomly selected flocks or herds periodically, using tests capable of distinguishing serological responses due to vaccination from those due to infection. These include complement fixation, immunodiffusion and competitive ELISA tests (active).
Active surveillance of tissues and blood samples from randomly selected animals of breeding age from randomly selected flocks and herds for bacteriology and serology.
Where the flock and individual animal prevalences are moderate to low and there are sufficient economic resources, a test and slaughter programme can be instituted. The conjunctival route of vaccination is compatible with this phase, as the bacteria are mainly restricted to the cranial lymph nodes, and although the immunity is similar to the subcutaneous route, the serological response is significantly reduced. Accordingly, when total eradication is the final objective, conjunctival vaccination of replacement animals with Rev.1 is the ideal tool for prophylaxis against B. melitensis infection in small ruminants.
In flocks or herds in which replacements have been continuously vaccinated for several years; there will obviously be a mixture of both infected and uninfected immune animals. Therefore the Rose Bengal or the indirect ELISA tests should be carried out periodically on all, or at least a representative random sample of, animals. Positive reactors can then be tested with the complement fixation, immunodiffusion or cELISA tests, and removed if positive.
Currently, there are no equivalents of the milk ring test for use in dairy animals. The brucellin test may also be used to screen unvaccinated flocks to determine their infection status.
Abortion incidents, movement tests, adjacent herds or flocks, and epidemiological investigation of infected flocks and herds can be monitored as in bovine brucellosis.
Because of their size and mobility, the marketing channels and slaughter locations of small ruminants, particularly in heavily infected regions, are so diverse that off-farm surveillance is problematical. If routes from to slaughter can be clearly defined, and trace-back possible, it may be practical to monitor long-terms trends by this method.
According to the OIE International Animal Health Code (Chapter 2.4.2 - Caprine and Ovine Brucellosis excluding B. ovis infections), for a country to be considered as officially free from brucellosis the following must apply:
For a country or part of the territory of a country to maintain its status as officially free from caprine and ovine brucellosis, a serological survey should be carried out every year on farms or in abattoirs on a representative sample of the caprine and ovine flocks of the country or part of the territory of the country sufficient to provide at least a 99% level of confidence of detecting caprine and ovine brucellosis if it is present at a prevalence rate exceeding 0.2% of the flocks.
For OIE purposes, a sheep flock or goat herd officially free from caprine and ovine brucellosis must satisfy the following requirements:
However, for flocks situated in a country or part of the territory of a country that qualifies as officially free under this paragraph, maintenance testing is not required.
For a flock to maintain its status as officially free from caprine and ovine brucellosis, a sample of the animals in the flock must be subjected each year to a diagnostic test for brucellosis, with negative results:
all males over six months of age;
all the animals introduced into the flock since the previous control; and
- 25% of the pubescent females; the number of females included in the sample should not be less than 50, unless the flock contains fewer than 50 females, in which case all pubescent females should be included.
Controls must be carried out at up to three-year intervals, if the flock is situated in a part of the territory of the country where 99% of flocks are officially free from caprine and ovine brucellosis and the remainder are submitted to an eradication programme.
However, for flocks situated in a country or part of the territory of a country qualified as officially free (as noted above) maintenance testing is not required.
Whatever the periodicity of controls and the ways the status has been obtained, sheep and goats must only be introduced into the flocks in accordance with the stated requirements.
If a sheep or goat reacts positively to a diagnostic test for caprine and ovine brucellosis, the status of flock officially free from brucellosis shall be suspended and may not be recovered unless the following requirements have been fulfilled:
|  Obtainable
from: Institut National de la Recherche Agronomique (INRA), Laboratoire
de pathologie infectieuse et immunologie, 37380 Nouzilly, France.|