Control of contagious bovine pleuropneumonia: knowledge, attitudes, perceptions and practices in Narok district of Kenya

CBPP is an important transboundary disease in sub-Saharan Africa whose control is urgent. Participatory data collection involving 52 focus group discussions in 37 village clusters and key informant interviews, a cross-sectional study involving 232 households and a post-vaccination follow up involving 203 households was carried out in 2006-2007 in Narok South district of Kenya. This was to investigate knowledge, attitudes, perceptions and practices (KAPP) associated with control of CBPP as well as the adverse post-vaccination reactions in animals in order to advice the control policy. The community perceived trans-boundary CBPP threat to their cattle. They had traditional disease coping mechanisms and were conversant with CBPP prevention and control with 49.8% (95%CI: 42.8-56.7%) giving priority to CBPP control. However, 12.9% (95%CI: 9.0-18.1%) of pastoralists had no knowledge of any prevention method and 10.0% (95%CI: 6.5-14.7%) would not know what to do or would do nothing in the event of an outbreak. Although 43.5% (95%CI: 37.1-50.2%) of pastoralists were treating CBPP cases with antimicrobials, 62.5% (95%CI: 52.1-71.7%) of them doubted the effectiveness of the treatments. Pastoralists perceived vaccination to be the solution to CBPP but vaccination was irregular due to unavailability of the vaccine. Vaccination was mainly to control outbreaks rather than preventive and exhibited adverse post-vaccination reactions among 70.4% (95%CI: 63.6-76.5%) of herds and 3.8% (95%CI: 3.5-4.2%) of animals. Consequently, nearly 25.2% (95%CI: 18.5-33.2%) of pastoralists may resist subsequent vaccinations against CBPP. Pastoralists preferred CBPP vaccination at certain times of the year and that it is combined with other vaccinations. In conclusion, pastoralists were not fully aware of the preventive measures and interventions and post-vaccination reactions may discourage subsequent CBPP vaccinations. Consequently there is need for monitoring and management of post vaccination reactions and awareness creation on CBPP prevention and interventions and their merits and demerits. CBPP vaccine was largely unavailable to the pastoralists and the preference of the pastoralists was for vaccination at specified times and vaccine combinations which makes it necessary to avail the vaccine in conformity with the pastoralists preferences. In addition, planning vaccinations should involve pastoralists and neighbouring countries. As the results cannot be generalized, further studies on CBPP control methods and their effectiveness are recommended.

Willingness to pay for contagious bovine pleuropneumonia vaccination in Narok South District of Kenya

Contagious bovine pleuropneumonia (CBPP) is an economically important trans-boundary cattle disease which affects food security and livelihoods. A conjoint analysis–contingent valuation was carried out on 190 households in Narok South District of Kenya to measure willingness to pay (WTP) and demand for CBPP vaccine and vaccination as well as factors affecting WTP. The mean WTP was calculated at Kenya Shillings (KSh) 212.48 (USD 3.03) for vaccination using a vaccine with the characteristics that were preferred by the farmers (preferred vaccine and vaccination) and KSh −71.45 (USD −1.02) for the currently used vaccine and vaccination. The proportion of farmers willing to pay an amount greater than zero was 66.7% and 34.4% for the preferred and current vaccine and vaccination respectively. About one third (33.3%) of farmers would need to be compensated an average amount of KSh 1162.62 (USD 13.68) per animal to allow their cattle to be vaccinated against CBPP using the preferred vaccine and vaccination. About two-thirds (65.6%) of farmers would need to be compensated an average amount of KSh 853.72 (USD 12.20) per animal to allow their cattle to be vaccinated against CBPP using the current vaccine and vaccination. The total amount of compensation would be KSh 61.39 million (USD 0.88 million) for the preferred vaccine and vaccination and KSh 90.15 million (USD 1.29 million) for the current vaccine and vaccination. Demand curves drawn from individual WTP demonstrated that only 59% and 27% of cattle owners with a WTP greater than zero were willing to pay a benchmark cost of KSh 34.60 for the preferred and current vaccine respectively. WTP was negatively influenced by the attitude about household economic situation (p = 0.0078), presence of cross breeds in the herd (p < 0.0001) and years since CBPP had been experienced in the herd (p = 0.0375). It was positively influenced by education (p = 0.0251) and the practice of treating against CBPP (p = 0.0432). The benefit cost ratio (BCR) for CBPP vaccination was 2.9–6.1 depending on the vaccination programme. In conclusion, although a proportion of farmers was willing to pay, participation levels may be lower than those required to interrupt transmission of CBPP. Households with characteristics that influence WTP negatively need persuasion to participate in CBPP vaccination. It is economically worthwhile to vaccinate against CBPP. A benefit cost analysis (BCA) using aggregated WTP as benefits can be used as an alternative method to the traditional BCA which uses avoided production losses (new revenue) and costs saved as benefits.

Exploring farmer preferences for contagious bovine pleuropneumonia vaccination: a case study of Narok District, Kenya

Contagious bovine pleuropneumonia (CBPP) is an economically important disease in most of sub-Saharan Africa. A conjoint analysis and ordered probit regression models were used to measure the preferences of farmers for CBPP vaccine and vaccination attributes. This was with regard to inclusion or not of an indicator in the vaccine, vaccine safety, vaccine stability as well as frequency of vaccination, vaccine administration and the nature of vaccination. The analysis was carried out in 190 households in Narok District of Kenya between October and December 2006 using structured questionnaires, 16 attribute profiles and a five-point Likert scale. The factors affecting attribute valuation were shown through a two-way location interaction model. The study also demonstrated the relative importance (RI) of attributes and the compensation value of attribute levels. The attribute coefficient estimates showed that farmers prefer a vaccine that has an indicator, is 100% safe and is administered by the government (p<0.0001). The preferences for the vaccine attributes were consistent with expectations. Preferences for stability, frequency of vaccination and nature of vaccination differed amongst farmers (p>0.05). While inclusion of an indicator in the vaccine was the most important attribute (RI=43.6%), price was the least important (RI=0.5%). Of the 22 household factors considered, 15 affected attribute valuation. The compensation values for a change from non inclusion to inclusion of an indicator, 95-100% safety, 2h to greater than 2h stability and from compulsory to elective vaccination were positive while those for a change from annual to biannual vaccination and from government to private administration were negative. The study concluded that the farmers in Narok District had preferences for specific vaccine and vaccination attributes. These preferences were conditioned by various household characteristics and disease risk factors. On average the farmers would need to be compensated or persuaded to accept biannual and private vaccination against CBPP. There is need for consideration of farmer preferences for vaccine attribute levels during vaccine formulations and farmer preferences for vaccination attribute levels when designing delivery of vaccines.

Contagious bovine pleuropneumonia and Mycoplasma mycoides var. mycoides, 1852-1968 : a bibliography and index ; appendix 1969-April 1970 /

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Isothermal loop-mediated amplification (LAMP) for diagnosis of contagious bovine pleuro-pneumonia.

BACKGROUND Contagious Bovine Pleuropneumonia (CBPP) is the most important chronic pulmonary disease of cattle on the African continent causing severe economic losses. The disease, caused by infection with Mycoplasma mycoides subsp. mycoides is transmitted by animal contact and develops slowly into a chronic form preventing an early clinical diagnosis. Because available vaccines confer a low protection rate and short-lived immunity, the rapid diagnosis of infected animals combined with traditional curbing measures is seen as the best way to control the disease. While traditional labour-intensive bacteriological methods for the detection of M. mycoides subsp. mycoides have been replaced by molecular genetic techniques in the last two decades, these latter approaches require well-equipped laboratories and specialized personnel for the diagnosis. This is a handicap in areas where CBPP is endemic and early diagnosis is essential. RESULTS We present a rapid, sensitive and specific diagnostic tool for M. mycoides subsp. mycoides detection based on isothermal loop-mediated amplification (LAMP) that is applicable to field conditions. The primer set developed is highly specific and sensitive enough to diagnose clinical cases without prior cultivation of the organism. The LAMP assay detects M. mycoides subsp. mycoides DNA directly from crude samples of pulmonary/pleural fluids and serum/plasma within an hour using a simple dilution protocol. A photometric detection of LAMP products allows the real-time visualisation of the amplification curve and the application of a melting curve/re-association analysis presents a means of quality assurance based on the predetermined strand-inherent temperature profile supporting the diagnosis. CONCLUSION The CBPP LAMP developed in a robust kit format can be run on a battery-driven mobile device to rapidly detect M. mycoides subsp. mycoides infections from clinical or post mortem samples. The stringent innate quality control allows a conclusive on-site diagnosis of CBPP such as during farm or slaughter house inspections.

Beta-D-glucoside utilization by Mycoplasma mycoides subsp. mycoides SC: possible involvement in the control of cytotoxicity towards bovine lung cells

BACKGROUND: Contagious bovine pleuropneumonia (CBPP) caused by Mycoplasma mycoides subsp. mycoides small-colony type (SC) is among the most serious threats for livestock producers in Africa. Glycerol metabolism-associated H2O2 production seems to play a crucial role in virulence of this mycoplasma. A wide number of attenuated strains of M. mycoides subsp. mycoides SC are currently used in Africa as live vaccines. Glycerol metabolism is not affected in these vaccine strains and therefore it does not seem to be the determinant of their attenuation. A non-synonymous single nucleotide polymorphism (SNP) in the bgl gene coding for the 6-phospho-beta-glucosidase (Bgl) has been described recently. The SNP differentiates virulent African strains isolated from outbreaks with severe CBPP, which express the Bgl isoform Val204, from strains to be considered less virulent isolated from CBPP outbreaks with low mortality and vaccine strains, which express the Bgl isoform Ala204. RESULTS: Strains of M. mycoides subsp. mycoides SC considered virulent and possessing the Bgl isoform Val204, but not strains with the Bgl isoform Ala204, do trigger elevated levels of damage to embryonic bovine lung (EBL) cells upon incubation with the disaccharides (i.e., beta-D-glucosides) sucrose and lactose. However, strains expressing the Bgl isoform Val204 show a lower hydrolysing activity on the chromogenic substrate p-nitrophenyl-beta-D-glucopyranoside (pNPbG) when compared to strains that possess the Bgl isoform Ala204. Defective activity of Bgl in M. mycoides subsp. mycoides SC does not lead to H2O2 production. Rather, the viability during addition of beta-D-glucosides in medium-free buffers is higher for strains harbouring the Bgl isoform Val204 than for those with the isoform Ala204. CONCLUSION: Our results indicate that the studied SNP in the bgl gene is one possible cause of the difference in bacterial virulence among strains of M. mycoides subsp. mycoides SC. Bgl does not act as a direct virulence factor, but strains possessing the Bgl isoform Val204 with low hydrolysing activity are more prone to survive in environments that contain high levels of beta-D-glucosides, thus contributing in some extent to mycoplasmaemia.

Mycoplasma leachii sp. nov. as a new species designation for Mycoplasma sp. bovine group 7 of Leach, and reclassification of Mycoplasma mycoides subsp. mycoides LC as a serovar of Mycoplasma mycoides subsp. capri

The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii.

Comparative analysis of four lipoproteins from Mycoplasma mycoides subsp. mycoides Small Colony identifies LppA as a major T-cell antigen

Control of contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), remains an important goal in Africa. Subunit vaccines triggering B and T-cell responses could represent a promising approach. To this aim, the T-cell immunogenicity of four MmmSC lipoproteins (LppA, LppB, LppC and LppQ), present in African strains and able to elicit humoral response, was evaluated. In vitro assays revealed that only LppA was recognized by lymph node lymphocytes taken from three cattle, 3 weeks after MmmSC exposure. Maintenance of the LppA-specific response, relying on CD4 T-cells and IFN gamma production, was then demonstrated 1 year after infection. LppA is thus an important target for the CD4 T-cells generated early after MmmSC infection and persisting in the lymph nodes of recovered cattle. Its role as a protective antigen and ability to in vivo trigger both arms of the host immune response remain to be evaluated.

Detection of Mycoplasma mycoides subsp. mycoides SC in bronchoalveolar lavage fluids of cows based on a TaqMan real-time PCR discriminating wild type strains from an lppQ(-) mutant vaccine strain used for DIVA-strategies

Contagious bovine pleuropneumonia (CBPP) is the most serious cattle disease in Africa, caused by Mycoplasma mycoides subsp. mycoides small-colony type (SC). CBPP control strategies currently rely on vaccination with a vaccine based on live attenuated strains of the organism. Recently, an lppQ(-) mutant of the existing vaccine strain T1/44 has been developed (Janis et al., 2008). This T1lppQ(-) mutant strain is devoid of lipoprotein LppQ, a potential virulence attribute of M. mycoides subsp. mycoides SC. It is designated as a potential live DIVA (Differentiating Infected from Vaccinated Animals) vaccine strain allowing both serological and etiological differentiation. The present paper reports on the validation of a control strategy for CBPP in cattle, whereby a TaqMan real-time PCR based on the lppQ gene has been developed for the direct detection of M. mycoides subsp. mycoides SC in ex vivo bronchoalveolar lavage fluids of cows and for the discrimination of wild type strains from the lppQ(-) mutant vaccine strain.

Molecular mechanisms of pathogenicity of Mycoplasma mycoides subsp. mycoides SC

Mycoplasma mycoides subsp. mycoides SC, the aetiological agent of contagious bovine pleuropneumonia (CBPP), is considered the most pathogenic of the Mycoplasma species. Its virulence is probably the result of a coordinated action of various components of an antigenically and functionally dynamic surface architecture. The different virulence attributes allow the pathogen to evade the host's immune defence, adhere tightly to the host cell surface, persist and disseminate in the host causing mycoplasmaemia, efficiently import energetically valuable nutrients present in the environment, and release and simultaneously translocate toxic metabolic pathway products to the host cell where they cause cytotoxic effects that are known to induce inflammatory processes and disease. This strategy enables the mycoplasma to exploit the minimal genetic information in its small genome, not only to fulfil the basic functions for its replication but also to damage host cells in intimate proximity thereby acquiring the necessary bio-molecules, such as amino acids and nucleic acid precursors, for its own biosynthesis and survival.

Domain analysis of lipoprotein LppQ in Mycoplasma mycoides subsp. mycoides SC

The lipoprotein LppQ is the most prominent antigen of Mycoplasma mycoides subsp. mycoides small colony type (SC) during infection of cattle. This pathogen causes contagious bovine pleuropneumonia (CBPP), a devastating disease of considerable socio-economic importance in many countries worldwide. The dominant antigenicity and high specificity for M. mycoides subsp. mycoides SC of lipoprotein LppQ have been exploited for serological diagnosis and for epidemiological investigations of CBPP. Scanning electron microscopy and immunogold labelling were used to provide ultrastructural evidence that LppQ is located to the cell membrane at the outer surface of M. mycoides subsp. mycoides SC. The selectivity and specificity of this method were demonstrated through discriminating localization of extracellular (i.e., in the zone of contact with host cells) vs. integral membrane domains of LppQ. Thus, our findings support the suggestion that the accessible N-terminal domain of LppQ is surface exposed and such surface localization may be implicated in the pathogenesis of CBPP.

Functional and antigenic properties of GlpO from Mycoplasma mycoides subsp. mycoides SC: characterization of a flavin adenine dinucleotide-binding site deletion mutant

L-alpha-glycerophosphate oxidase (GlpO) plays a central role in virulence of Mycoplasma mycoides subsp. mycoides SC, a severe bacterial pathogen causing contagious bovine pleuropneumonia (CBPP). It is involved in production and translocation of toxic H(2)O(2) into the host cell, causing inflammation and cell death. The binding site on GlpO for the cofactor flavin adenine dinucleotide (FAD) has been identified as Gly(12)-Gly(13)-Gly(14)-Ile(15)-Ile(16)-Gly(17). Recombinant GlpO lacking these six amino acids (GlpODeltaFAD) was unable to bind FAD and was also devoid of glycerophosphate oxidase activity, in contrast to non-modified recombinant GlpO that binds FAD and is enzymatically active. Polyclonal monospecific antibodies directed against GlpODeltaFAD, similarly to anti-GlpO antibodies, neutralised H(2)O(2) production of M. mycoides subsp. mycoides SC grown in the presence of glycerol, as well as cytotoxicity towards embryonic calf nasal epithelial (ECaNEp) cells. The FAD-binding site of GlpO is therefore suggested as a valuable target site for the future construction of deletion mutants to yield attenuated live vaccines of M. mycoides subsp. mycoides SC necessary to efficiently combat CBPP.

Cattle immunized against the pathogenic L-α-glycerol-3-phosphate oxidase of Mycoplasma mycoides subs. mycoides fail to generate neutralizing antibodies and succumb to disease on challenge

The membrane-associated enzyme L-α-glycerol-3-phosphate oxidase (GlpO) of Mycoplasma mycoides subs. mycoides (Mmm), the causal agent of contagious bovine pleuropneumonia (CBPP) has been identified as a virulence factor responsible for the release of toxic by-products such as H2O2 that mediate host cell injury. Since CBPP pathogenesis is based on host inflammatory reactions, we have determined the capacity of recombinant GlpO to generate in vivo protective responses against challenge in immunized cattle. We also investigated whether sera raised against recombinant GlpO in cattle and mice inhibit production of H2O2 by Mmm. Immunization of cattle with recombinant GlpO did not protect against challenge with a virulent strain of Mmm. Further, although both murine and bovine antisera raised against recombinant GlpO detected recombinant and native forms of GlpO in immunoblot assays with similar titres, only murine antibodies could neutralize GlpO enzymatic function. The data raise the possibility that Mmm has adapted to evade potential detrimental antibody responses in its definitive host.

The origin of the 'Mycoplasma mycoides cluster' coincides with domestication of ruminants.

The 'Mycoplasma mycoides cluster' comprises the ruminant pathogens Mycoplasma mycoides subsp. mycoides the causative agent of contagious bovine pleuropneumonia (CBPP), Mycoplasma capricolum subsp. capripneumoniae the agent of contagious caprine pleuropneumonia (CCPP), Mycoplasma capricolum subsp. capricolum, Mycoplasma leachii and Mycoplasma mycoides subsp. capri. CBPP and CCPP are major livestock diseases and impact the agricultural sector especially in developing countries through reduced food-supply and international trade restrictions. In addition, these diseases are a threat to disease-free countries. We used a multilocus sequence typing (MLST) approach to gain insights into the demographic history of and phylogenetic relationships among the members of the 'M. mycoides cluster'. We collected partial sequences from seven housekeeping genes representing a total of 3,816 base pairs from 118 strains within this cluster, and five strains isolated from wild Caprinae. Strikingly, the origin of the 'M. mycoides cluster' dates to about 10,000 years ago, suggesting that the establishment and spread of the cluster coincided with livestock domestication. In addition, we show that hybridization and recombination may be important factors in the evolutionary history of the cluster.