Differential diagnosis of tick-borne diseases and population genetic analysis of Babesia bovis and Babesia bigemina

Abstract: Tick-borne diseases are a constraint to livestock production in many developing countries. They are responsible for high morbidity and mortality resulting in decreased production of meat, milk and other livestock by-products. The most important tick-borne diseases of livestock in sub-Saharan Africa are East coast fever (caused by Theileria parva), babesiosis (caused by Babesia bigemina and Ba. bovis), anaplasmosis (caused by Anaplasma marginale) and heartwater (caused by Ehrlichia ruminantium). Despite their economic importance, information on the epidemiology of these diseases in many countries is often lacking or inadequate, resulting in inappropriate disease control strategies being implemented. The availability of specific, sensitive and cost-effective diagnostic methods is important in the design and implementation of effective disease control strategies. In this study PCR assays based on the 18S and 16S rRNA gene sequences, that could identify Theileria / Babesia and Anaplasma / Ehrlichia pathogens of cattle respectively, were developed. In addition, PCR assays based on the β-tubulin gene that could detect T. parva, Ba. bigemina, Ba. bovis and T. taurotragi, and PCR assays based on the cytochrome b gene that could diagnose infection by Ba. bigemina and Ba. bovis were also developed. When the 18S and 16S rRNA gene PCR assays were combined into a multiplex PCR assay, Ba. bigemina and E. ruminantium DNA did not amplify and some non-specific bands were observed following agarose gel electrophoresis. The β-tubulin gene multiplex PCR assay for the diagnosis of T. parva, Ba. bovis and Ba. bigemina worked relatively well when used on laboratory-derived parasite DNA preparations. However, when it was used on field samples collected on FTA cards, multiple non-specific bands were observed after agarose gel electrophoresis of the PCR products. The 18S and 16S rRNA gene PCR assays were used for an epidemiological study of tick-borne diseases of cattle in Central and Eastern Zambia in the wet and dry seasons. All the disease pathogens under study (T. parva, T. mutans, T. taurotragi, Ba. bovis, Ba. bigemina, Anaplasma spp and E. ruminantium) were prevalent in all the regions of the country in both seasons. However, variation was observed in the prevalence of these pathogens between the regions and the seasons. A number of risk factors, associated with the occurrence of tick-borne pathogens in cattle and the tick burdens observed on cattle in the wet season were determined. A negative association was observed between the number of co-infecting pathogens and the erythrocyte packed cell volume (PCV) of carrier cattle. Using recently available genome sequences, mini- and microsatellite markers were developed for population genetic analysis of Ba. bovis and Ba. bigemina parasite populations. Ba. bovis isolates from Zambia and Turkey and Ba. bigemina isolates from Zambia were used in the population genetic analysis. High levels of genetic diversity were observed for both parasites. Population genetic analysis of the Zambian and Turkish Ba. bovis populations, using eight genetic markers showed that the two populations were sub-structured. The Zambian population comprised a single randomly mating population, while the Turkish population comprised two genetically distinct subpopulations. Population genetic analysis of the Ba. bigemina parasites from Zambia showed that this parasite population was in linkage disequilibrium. Further, analysis of the Ba. bigemina population using STRUCTURE showed that it was genetically sub-structured into five distinct subgroups. However, the resulting sample size of each subgroup was too small to definitely determine whether they were panmictic. These results provide an improved understanding of the epidemiology of bovine Babesia parasites in Turkey and Zambia

Policy Concerns, Opportunities, Challenges, and Attitude towards One Health Practice in Zambia

One Health in terms of collaboration, particularly between human and animal health sectors to prevent and control zoonoses has been low while the sectors have a lot of things in common. Such common things include aspects of disease causative agents (viruses, bacteria, parasites, etc.) and those of disease occurrence mediator conditions (social, cultural, economic or climatic). Therefore, the research from which this paper is based was done with the objectives to: (a) assess the extent to which human and animal health policies facilitate one health in terms of collaboration; (b) rank opportunities for and challenges to collaboration among medical, and veterinary officers according to the views and experiences of the respondents in the Ministry of Health and Ministry of Agriculture; and (c) determine the attitude of the respondents towards One Health approaches in terms of collaboration in dealing with zoonoses. A cross-sectional research design was used in this study whereby data were collected at a single point in time without repetition. Purposive sampling method was used to make sure that the respondents were only officials who usually participated in policy formulation in the two Ministries. It was found that almost three quarters (73.1%) of the respondents from both ministries agreed that there was no policy which directly facilitated One Health in terms of collaboration. It was also found that 83.6% of the respondents pointed out that human and animal health policy making process was a top-down process. Furthermore, it was found that the main opportunities that could enhance collaboration were sufficient money in budgeting; advocacy for control of neglected zoonotic diseases in human and animal health; and one health policy formulation (71.3%, 68.2% and 65.5% respectively). The overall attitude towards collaboration among respondents was favourable; they scored an average of 62.2 out of 100.0 points on a Likert scale. It is concluded that if opportunities enhancing collaboration were strengthened and challenges to collaboration were overcome, human health and animal health experts could collaborate more in reduction of disease burden in both humans and livestock. Keywords: One health, policy, attitude, opportunities, challenge

Documenting the absence of bovine brucellosis in dairy cattle herds in the southern region of Malawi and the associated knowledge, attitudes and practices of farmers

Source at https://www.jsava.co.za/index.php/jsava/article/view/473.There is paucity of Brucella prevalence data in Malawi. For this reason, a cross-sectional study was conducted, from 06 January 2020 to 27 February 2020, to estimate the seroprevalence of brucellosis in dairy cattle herds amongst smallholder farmers, government and private dairy farms in the southern region. A total of 529 serum samples were screened for anti-Brucella antibodies using the Rose Bengal test (RBT) and a competitive enzyme-linked immunosorbent assay (cELISA). A pre-tested electronic (Epicollect tool, Wellcome Sanger Institute, United Kingdom) questionnaire was administered to 378 smallholder farmers to assess their knowledge, attitudes and practices towards brucellosis. Descriptive statistics were used to analyse the data in Microsoft Excel® and Statistical Package for Social Sciences (SPSS®) version 21. No animal tested positive for presence of anti-Brucella antibodies, indicating 0% prevalence (individual and herd levels). The majority (94.2%; 95% confidence interval [CI]: 91.8–96.5) of smallholder farmers had never heard about brucellosis. Furthermore, assisting during parturition without protective equipment (41.3%; 95% CI: 36.3–46.2) and using bulls for breeding (75%; 95% CI: 70.2–78.9) were amongst the common risk practices that were identified. We could not detect brucellosis in this study that indicates the disease could be very rare or even absent in the dairy cattle herds of the southern region of Malawi. However, further Brucella studies need to be conducted in cattle, small livestock, wildlife and humans to document the true status of brucellosis in the country. Brucellosis surveillance, monitoring, awareness and preventive measures are required to maintain this favourable situation. Keywords: bovine brucellosis (contagious abortion); dairy cattle herds; seroprevalence; knowledge; attitudes and practices; Malawi

Development of a multiplex PCR assay for simultaneous detection of Theileria annulata, Babesia bovis and Anaplasma marginale in cattle

Tropical theileriosis, bovine babesiosis and anaplasmosis are tick-borne protozoan diseases that impose serious constraints on the health and productivity of domestic cattle in tropical and sub-tropical regions of the world. A common feature of these diseases is that, following recovery from primary infection, animals become persistent carriers of the pathogen and continue to play a critical role in disease epidemiology, acting as reservoirs of infection. This study describes development and evaluation of multiplex and single PCR assays for simultaneous detection of Theileria annulata, Babesia bovis and Anaplasma marginale in cattle. Following in silico screening for candidate target genes representing each of the pathogens, an optimised multiplex PCR assay was established using three primer sets, cytob1, MAR1bB2 and bovar2A, for amplification of genomic DNA of T. annulata, A. marginale and B. bovis respectively. The designed primer sets were found to be species-specific, generating amplicons of 312, 265 and 166 base pairs, respectively and were deemed suitable for the development of a multiplex assay. The sensitivity of each primer pair was evaluated using serial dilutions of parasite DNA, while specificity was confirmed by testing for amplification from DNA of different stocks of each pathogen and other Theileria, Babesia and Anaplasma species. Additionally, DNA preparations derived from field samples were used to evaluate the utility of the single and multiplex PCRs for determination of infection status. The multiplex PCR was found to detect each pathogen species with the same level of sensitivity, irrespective of whether its DNA was amplified in isolation or together with DNA representing the other pathogens. Moreover, single and multiplex PCRs were able to detect each species with equal sensitivity in serially diluted DNA representing mixtures of T. annulata, B. bovis and A. marginale, and no evidence of non-specific amplification from non-target species was observed. Validation that the multiplex PCR efficiently detects single and mixed infections from field samples was demonstrated. The developed assay represents a simple and efficient diagnostic for co-detection of tropical theileriosis, bovine babesiosis and anaplasmosis, and may be a valuable tool for epidemiological studies aimed at assessing the burden of multiple infection with tick-borne pathogens and improving control of the associated diseases in endemic regions

Antibiogram bakterije Escherichia coli i enterokoka izdvojenih iz goveđega izmeta na području Kafue u Zambiji

Antimicrobial resistance to a panel of ten agents was determined by the disc diffusion technique for 83 Escherichia coli isolates, 29 Enterococcus faecium isolates and 62 Enterococcus faecalis isolates from faecal samples of apparently healthy pastoral cattle in the wildlife/livestock interface areas. Of all the E. coli isolates, 8% were diarrhoeagenic E. coli strains, 6% were enteropathogenic E. coli strains and 2% were enterotoxigenic E. coli strains. A high frequency of E. coli resistance to penicillin, erythromycin, cotrimoxazole and nitrofurantoin was observed. Enterococci showed the highest percentage of resistance to gentamycin, amoxycillin, ampicillin and tetracycline. None of the E. coli strains and Enterococci strains was resistant to tetracycline and vancomycin respectively. The results of this study underscore the presence of an animal reservoir of antibiotic resistant microorganisms that have the potential to enter the food chain.Osjetljivost 83 izolata vrste Escherichia coli, 29 izolata vrste Enterococcus faecium i 62 izolata vrste Enterococcus faecalis izdvojenih iz uzoraka izmeta klinički zdravih goveda u području dodira domaćih i divljih životinja, određena je difuzijskim postupkom prema deset antimikrobnih pripravaka. Od ukupno izdvojenih, 8% sojeva bakterije E. coli bilo je dijarejogeno, 6% enteropatogeno i 2% enterotoksigeno. Ustanovljena je česta otpornost vrste E. coli prema penicilinu, eritromicinu, kotrimoksazolu i nitrofurantoinu. Enterokoki su u najvećem postotku bili otporni prema gentamicinu, amoksicilinu, ampicilinu i tetraciklinu. Nijedan od izolata E. coli nije bio otporan prema tetraciklinu, a nijedan od enterokoka nije bio otporan prema vankomicinu. Rezultati istraživanja upućuju na postojanje životinjskoga rezervoara bakterija otpornih na antibiotike za koje postoji mogućnost da se prenesu u ljudski prehrambeni lanac

Antibiogram bakterije Escherichia coli i enterokoka izdvojenih iz goveđega izmeta na području Kafue u Zambiji

Antimicrobial resistance to a panel of ten agents was determined by the disc diffusion technique for 83 Escherichia coli isolates, 29 Enterococcus faecium isolates and 62 Enterococcus faecalis isolates from faecal samples of apparently healthy pastoral cattle in the wildlife/livestock interface areas. Of all the E. coli isolates, 8% were diarrhoeagenic E. coli strains, 6% were enteropathogenic E. coli strains and 2% were enterotoxigenic E. coli strains. A high frequency of E. coli resistance to penicillin, erythromycin, cotrimoxazole and nitrofurantoin was observed. Enterococci showed the highest percentage of resistance to gentamycin, amoxycillin, ampicillin and tetracycline. None of the E. coli strains and Enterococci strains was resistant to tetracycline and vancomycin respectively. The results of this study underscore the presence of an animal reservoir of antibiotic resistant microorganisms that have the potential to enter the food chain.Osjetljivost 83 izolata vrste Escherichia coli, 29 izolata vrste Enterococcus faecium i 62 izolata vrste Enterococcus faecalis izdvojenih iz uzoraka izmeta klinički zdravih goveda u području dodira domaćih i divljih životinja, određena je difuzijskim postupkom prema deset antimikrobnih pripravaka. Od ukupno izdvojenih, 8% sojeva bakterije E. coli bilo je dijarejogeno, 6% enteropatogeno i 2% enterotoksigeno. Ustanovljena je česta otpornost vrste E. coli prema penicilinu, eritromicinu, kotrimoksazolu i nitrofurantoinu. Enterokoki su u najvećem postotku bili otporni prema gentamicinu, amoksicilinu, ampicilinu i tetraciklinu. Nijedan od izolata E. coli nije bio otporan prema tetraciklinu, a nijedan od enterokoka nije bio otporan prema vankomicinu. Rezultati istraživanja upućuju na postojanje životinjskoga rezervoara bakterija otpornih na antibiotike za koje postoji mogućnost da se prenesu u ljudski prehrambeni lanac

A polymorphism in the haptoglobin, haptoglobin related protein locus is associated with risk of human sleeping sickness within Cameroonian populations

Human African Trypanosomiasis (HAT) is a neglected disease targeted for elimination as a public health problem by 2020. Elimination requires a better understanding of the epidemiology and clinical evolution of HAT. In addition to the classical clinical evolution of HAT, asymptomatic carriers and spontaneous cure have been reported in West Africa. A genetic component to human susceptibility to HAT has been suggested to explain these newly observed responses to infection. In order to test for genetic associations with infection response, genetic polymorphism in 17 genes were tested (APOL1, IL1B, IL4, IL4R, IL6, IL8, IL12B, IL12RB1, IL10, TNFA, INFG, MIF, HLA-G, HLA-A, HP, HPR and CFH). A case-control study was performed on 180 blood samples collected from 56 cases and 124 controls from Cameroon. DNA was extracted from blood samples. After quality control, 25 samples (24 controls and 1 case) were eliminated. The genotyping undertaken on 155 individuals including 55 cases and 100 controls were investigated at 96 loci (88 SNPs and 8 indels) located on 17 genes. Associations between these loci and HAT were estimated via a case-control association test. Analyses of 64 SNPs and 4 indels out of 96 identified in the selected genes reveal that the minor allele (T) of rs8062041 in haptoglobin (HP) appeared to be protective against HAT (p = 0.0002395, OR 0.359 (CI95 [0.204-0.6319])); indicating higher frequency in cases compared to controls. This minor allele with adjusted p value of 0.0163 is associated with a lower risk (protective effect) of developing sleeping sickness. The haptoglobin related protein HPR and HP are tightly linked and both are duplicated in some people and may lead to higher activity. This increased production could be responsible of the protection associated with rs8062041 even though this SNP is within HP

Exploring the Effect of Human and Animal Population Growth on Vector-Borne Disease Transmission with an Agent-Based Model of Rhodesian Human African Trypanosomiasis in Eastern Province, Zambia

This paper presents the development of an agent-based model (ABM) to investigate Trypanosoma brucei rhodesiense human African trypanosomiasis (rHAT) disease transmission. The ABM model, fitted at a fine spatial scale, was used to explore the impact of a growing host population on the spread of disease along a 75 km transect in the Luangwa Valley, Zambia. The model was used to gain a greater understanding of how increases in human and domestic animal population could impact the contact network between vector and host, the subsequent transmission patterns, and disease incidence outcomes in the region. Modelled incidence rates showed increases in rHAT transmission in both humans and cattle. The primary demographic attribution of infection switched dramatically from young children of both sexes attending school, to adult women performing activities with shorter but more frequent trips, such as water and firewood collection, with men more protected due to the presence of cattle in their routines. The interpretation of model output provides a plausible insight into both population development and disease transmission in the near future in the region and such techniques could aid well-targeted mitigation strategies in the future

Exploring the Effect of Human and Animal Population Growth on Vector-Borne Disease Transmission with an Agent-Based Model of Rhodesian Human African Trypanosomiasis in Eastern Province, Zambia

This paper presents the development of an agent-based model (ABM) to investigate Trypanosoma brucei rhodesiense human African trypanosomiasis (rHAT) disease transmission. The ABM model, fitted at a fine spatial scale, was used to explore the impact of a growing host population on the spread of disease along a 75 km transect in the Luangwa Valley, Zambia. The model was used to gain a greater understanding of how increases in human and domestic animal population could impact the contact network between vector and host, the subsequent transmission patterns, and disease incidence outcomes in the region. Modelled incidence rates showed increases in rHAT transmission in both humans and cattle. The primary demographic attribution of infection switched dramatically from young children of both sexes attending school, to adult women performing activities with shorter but more frequent trips, such as water and firewood collection, with men more protected due to the presence of cattle in their routines. The interpretation of model output provides a plausible insight into both population development and disease transmission in the near future in the region and such techniques could aid well-targeted mitigation strategies in the future