High co-infection rates of Babesia bovis, Babesia bigemina, and Anaplasma marginale in water buffalo in Western Cuba

  • Dasiel ObregónEmail author
  • Alejandro Cabezas-Cruz
  • Yasmani Armas
  • Jenevaldo B. Silva
  • Adivaldo H. Fonseca
  • Marcos R. André
  • Pastor Alfonso
  • Márcia C.S. Oliveira
  • Rosangela Z. Machado
  • Belkis Corona-González
Protozoology - Original Paper


Water buffalo is important livestock in several countries in the Latin American and Caribbean regions. This buffalo species can be infected by tick-borne hemoparasites and remains a carrier of these pathogens which represent a risk of infection for more susceptible species like cattle. Therefore, studies on the epidemiology of tick-borne hemoparasites in buffaloes are required. In this study, the prevalence of Babesia bovis, Babesia bigemina, and Anaplasma marginale were determined in water buffalo herds of western Cuba. To this aim, a cross-sectional study covering farms with large buffalo populations in the region was performed. Eight buffalo herds were randomly selected, and blood samples were collected from 328 animals, including 63 calves (3–14 months), 75 young animals (3–5 years), and 190 adult animals (> 5 years). Species-specific nested PCR and indirect ELISA assays were used to determine the molecular and serological prevalences of each hemoparasite, respectively. The molecular and serological prevalence was greater than 50% for the three hemoparasites. Differences were found in infection prevalence among buffalo herds, suggesting that local epidemiological factors may influence infection risk. Animals of all age groups were infected, with a higher molecular prevalence of B. bigemina and A. marginale in young buffalo and calves, respectively, while a stepwise increase in seroprevalence of B. bovis and B. bigemina from calves to adult buffaloes was found. The co-infection by the three pathogens was found in 12% of animals, and when analyzed by pair, the co-infections of B. bovis and B. bigemina, B. bigemina and A. marginale, and B. bovis and A. marginale were found in 20%, 24%, and 26%, respectively, underlying the positive interaction between these pathogens infecting buffaloes. These results provide evidence that tick-borne pathogen infections can be widespread among water buffalo populations in tropical livestock ecosystems. Further studies should evaluate whether these pathogens affect the health status and productive performance of water buffalo and infection risk of these pathogens in cattle cohabiting with buffalo.


Water buffalo Tick-borne pathogens Prevalence Co-infections nPCR iELISA 



This work was supported by funding from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq-474648/210-9), Brazil; Empresa Brasileira de Pesquisa Agropecuária (Embrapa Pecuária Sudeste); Fundacão de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Brazil (Process 2012/21371-4); Coordination for the Improvement of Higher Education Personnel (Capes/MES- Projetos. Process 089110); and National Priority Program for Animal and Plant Health, Ministry of Agriculture (MINAGRI-P131LH003007), Cuba. Dasiel Obregon wishes to acknowledge the Coordination for the Improvement of Higher Education Personnel (CAPES), Brazil, for his sandwich Ph.D. grant (Capes/MES- Docentes, 2013). We are grateful to Leonhard Schnittger, PhD. for their exhaustive review, which greatly improved this paper.

Compliance with ethical standards

The procedures involving animals in this work were according to the principles established by The International Guiding Principles for Biomedical Research Involving Animals (2012). Consequently, the committee on ethics and animal welfare at CENSA approved the experimental design of this research. The Frontier Veterinary Service of the Institute of Veterinary Medicine under the Ministry of Agriculture of the Republic of Cuba authorized the export of the DNA samples used in this work, under the Zoosanitary Export Certification number R.S.1522010.

Conflicts of interest

The authors declare that they have no conflict of interest.

Supplementary material

436_2018_6194_Fig5_ESM.png (254 kb)
Supplementary Fig SF1

Limit of detection (LOD) of nPCR assays for molecular diagnosis of B. bovis (A), B. bigemina (B), and A. marginale (C). Ten-fold serial dilutions of DNA positive controls were used as template in the first round PCR, no-template reactions were used as negative controls. Lanes 1–7 correspond to the quantities of DNA 10 to 10−6 ng, respectively and M: DNA ladder. The LOD of nPCR assays of B. bovis and B. bigemina were established at 10−5 ng of DNA template, however, the LOD of the nPCR assay for A. marginale was in 10−6 ng. The size (base pair, bp) of the generated bands is indicated. (PNG 254 kb)

436_2018_6194_MOESM1_ESM.tif (1.3 mb)
High resolution image (TIF 1296 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Dasiel Obregón
    • 1
    • 2
    Email author
  • Alejandro Cabezas-Cruz
    • 3
  • Yasmani Armas
    • 1
  • Jenevaldo B. Silva
    • 4
  • Adivaldo H. Fonseca
    • 4
  • Marcos R. André
    • 5
  • Pastor Alfonso
    • 2
  • Márcia C.S. Oliveira
    • 6
  • Rosangela Z. Machado
    • 5
  • Belkis Corona-González
    • 2
  1. 1.Universidad Agraria de La HabanaSan José de Las LajasCuba
  2. 2.Centro Nacional de Sanidad AgropecuariaSan José de Las LajasCuba
  3. 3.UMR BIPAR, INRA, ANSES, Ecole Nationale Vétérinaire d’AlfortUniversité Paris-EstMaisons-AlfortFrance
  4. 4.Universidade Federal Rural do Rio de JaneiroSeropédica - RJBrazil
  5. 5.Universidade Estadual Paulista-Campus de JaboticabalJaboticabalBrazil
  6. 6.Embrapa Pecuária SudesteSão CarlosBrazil

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