The tick borne Babesia parasites remain an important limitation for development of cattle industries worldwide. A stable transfection of Babesia bovis will be useful for functional analysis of the recently sequenced B. bovis genome and to design improved methods to control Babesia infections. In thi...
Cao, Shinuo; Aboge, Gabriel Oluga; Terkawi, Mohamad Alaa; Yu, Longzheng; Kamyingkird, Ketsarin; Luo, Yuzi; Li, Yan; Goo, Youn-Kyoung; Yamagishi, Junya; Nishikawa, Yoshifumi; Yokoyama, Naoaki; Suzuki, Hiroshi; Igarashi, Ikuo; Maeda, Ryuichiro; Inpankaew, Tawin; Jittapalapong, Sathaporn; Xuan, Xuenan
Although Babesia bovis and Babesia bigemina infections cause economic losses in the cattle industry in northern Thailand, there is inadequate information on Babesia isolates present in the area. Therefore, to determine the prevalence and genetic relationship between Babesia isolates, we screened 200 blood samples of cattle from Chiang Rai, Chiang Mai, and Lumpang provinces of northern Thailand. A nested polymerase chain reaction using primers targeting B. bovis spherical body protein 2 (BboSBP2) and B. bigemina rhoptry-associated protein 1a (BbiRAP-1a) genes revealed a prevalence of 12 and 21 % for B. bovis and B. bigemina, respectively, while that of mixed infections was 6.5 % samples. The prevalences of B. bovis in Chiang Rai, Chiang Mai, and Lumpang were 9.5, 3.7, and 25.5 %, respectively. For B. bigemina, the prevalences were 15.8, 12.9, and 39.2 % in Chiang Rai, Chiang Mai, and Lumpang, respectively. Mixed infections with B. bovis and B. bigemina were 6.3 % in Chiang Rai, 1.9 % in Chiang Mai, and 13.7 % in Lumpang. The identical sequences of either BboSBP2 gene or BbiRAP-1a gene were shared among the Babesia isolates in the three provinces of northern Thailand. Further analysis using the internal transcribed spacer gene revealed at least four genotypes for B. bovis and five genotypes for B. bigemina in northern Thailand, while the sequences present great genetic diversities in the different isolates. Overall, we have demonstrated a high prevalence and polymorphism of Babesia parasites in northern Thailand calling for the need to design effective control programs for bovine babesiosis.
An immunosensor method for diagnosis of Babesia bovis in cattle based on impedance measurement is presented in this study. The method probes the interaction between serum antibodies against B. bovis infected cattle and recombinant protein, RAP-1, with C-terminal obtained from a Portuguese B. bovis s...
Maeda, Hiroki; Boldbaatar, Damdinsuren; Kusakisako, Kodai; Galay, Remil Linggatong; Aung, Kyaw Min; Umemiya-Shirafuji, Rika; Mochizuki, Masami; Fujisaki, Kozo; Tanaka, Tetsuya
Haemaphysalis longicornis is known as one of the most important ticks transmitting Babesia parasites in East Asian countries, including Babesia ovata and Babesia gibsoni, as well as Theileria parasites. H. longicornis is not the natural vector of Babesia bovis and Babesia bigemina. Vector ticks and transmitted parasites are thought to have established unique host-parasite interaction for their survival, meaning that vector ticks may have defensive molecules for the growth control of parasites in their bodies. However, the precise adaptation mechanism of tick-Babesia parasites is still unknown. Recently, cyclophilin A (CyPA) was reported to be important for the development of Babesia parasites in ticks. To reveal a part of their adaptation mechanism, the current study was conducted. An injection of B. bovis-infected RBCs into adult female H. longicornis ticks was found to upregulate the expression profiles of the gene and protein of CyPA in H. longicornis (HlCyPA). In addition, recombinant HlCyPA (rHlCyPA) purified from Escherichia coli exhibited significant inhibitory growth effects on B. bovis and B. bigemina cultivated in vitro, without any hemolytic effect on bovine RBCs at all concentrations used. In conclusion, our results suggest that HlCyPA might play an important role in the growth regulation of Babesia parasites in H. longicornis ticks, during natural acquisition from an infected host. Furthermore, rHlCyPA may be a potential alternative chemotherapeutic agent against babesiosis.
Aboge, Gabriel Oluga; Cao, Shinuo; Terkawi, Mohamad Alaa; Masatani, Tatsunori; Goo, Younkyoung; AbouLaila, Mahmoud; Nishikawa, Yoshifumi; Igarashi, Ikuo; Suzuki, Hiroshi; Xuan, Xuenan
The M17 leucine aminopeptidase (M17LAP) enzymes of the other apicomplexan parasites have been characterized and shown to be inhibited by bestatin. Though Babesia bovis also belongs to the apicomplexan group, it is not known whether its M17LAP could display similar biochemical properties as well as inhibition profile. To unravel this uncertainty, a B. bovis M17LAP (BbM17LAP) gene was expressed in Escherichia coli , and activity of the recombinant enzyme as well as its inhibition by bestatin were evaluated. The inhibitory effect of the compound on growths of B. bovis and Babesia gibsoni in vitro was also determined. The expression of the gene fused with glutathione S-transferase (GST) yielded approximately 81-kDa recombinant BbM17LAP (rBbM17LAP). On probing with mouse anti-rBbM17LAP serum, a green fluorescence was observed on the parasite cytosol on confocal laser microscopy, and a specific band greater than the predicted molecular mass was seen on Western blotting. The Km and Vmax values of the recombinant enzyme were 139.3 ± 30.25 and 64.83 ± 4.6 μM, respectively, while the Ki was 2210 ± 358 μM after the inhibition. Bestatin was a more potent inhibitor of the growth of B. bovis [IC50 (50% inhibition concentration) = 131.7 ± 51.43 μM] than B. gibsoni [IC50 = 460.8 ± 114.45 μM] in vitro. The modest inhibition of both the rBbM17LAP activity and Babesia parasites' growth in vitro suggests that this inhibition may involve the endogenous enzyme in live parasites. Therefore, BbM17LAP may be a target of bestatin, though more studies with other aminopeptidase inhibitors are required to confirm this.
Babesia bovis, an intra-erythrocytic tick-borne apicomplexan protozoan, is one of the agents of bovine babesiosis. Its life cycle includes sexual reproduction within cattle fever ticks, Rhipicephalus spp. Six B. bovis 6-Cys gene superfamily members were previously identified (A, B, C, D, E, F) and t...
Simuunza, Martin; Bilgic, Huseyin; Karagenc, Tulin; Syakalima, Michelo; Shiels, Brian; Tait, Andy; Weir, William
The tick-borne protozoan parasite, Babesia bovis is one of the causes of bovine babesiosis, an economically important disease of cattle in tropical and sub-tropical countries. Using the recently published genome sequence of the parasite, we developed a panel of eight mini- and micro-satellite markers and used these to investigate the role of genetic exchange in the population structure and diversity of the parasite using isolates from Zambia and Turkey. This population genetic analysis showed that genetic exchange occurs and that there are high levels of genetic diversity, with geographical sub-structuring quantified using Wright's F Index. Linkage disequilibrium was observed when isolates from both countries were treated as one population, but when isolates from Zambia were analysed separately linkage equilibrium was observed. The Turkish isolates were sub-structured, containing two genetically distinct sub-groups, both of which appeared to be in linkage equilibrium. The results of the Zambian study suggest that a sub-set of the parasite population is responsible for the westward spread of babesiosis into the previously disease-free central region of the country. The Zambian isolates had a significantly higher number of genotypes per sample than those from Turkey and age was found to be a significant predictor of the multiplicity of infection. The high levels of diversity seen in the Zambian and Turkish B. bovis populations have implications in the development of subunit vaccines against the disease and the spread of drug resistance.
The Rhipicephalus microplus tick is a notorious blood-feeding ectoparasite of livestock, especially cattle, responsible for massive losses in animal production. It is the main vector for transmission of pathogenic bacteria and parasites, including Babesia bovis, an intraerythrocytic apicomplexan pro...
A gene coding for a protein with sequence similarity to the Toxoplasma gondii micronemal 1 (MIC1) protein that contains a copy of a domain described as a sialic acid-binding micronemal adhesive repeat was identified in the Babesia bovis genome. The single copy gene, located in chromosome 3, contains...
Glycosylphosphatidyl-anchored proteins are particularly abundant on the surface of pathogenic protozoans and might play an important role for parasite survival. In the present work the relevance of GPI-anchored proteins for erythrocyte invasion of Babesia bovis, one of the tick-transmitted causative...
Boophilus ticks are vectors of Babesia bovis, the protozoan causative agent of cattle fever, a disease which is responsible for significant production losses to cattle producers in much of Africa, Central and South America and Australia. We utilized subtractive cDNA library synthesis techniques to o...
Mini and microsatellite sequences have proven to be excellent tools for the differentiation of strains and populations in several protozoan parasites due to their high variability. In the present work we have searched the genome of the tick-transmitted bovine hemoprotozoon Babesia bovis for tandem r...
Wildlife are an important component in the vector-host-pathogen triangle of livestock diseases, as they maintain biological vectors that transmit pathogens and can serve as reservoirs for such infectious pathogens. Babesia bovis is a tick-borne pathogen, vectored by cattle fever ticks, Rhipicephalus...
Babesia bovis is an apicomplexan tick-transmitted pathogen of cattle imposing a global risk and severe constraints to livestock health and economic development. The complete genome sequence was undertaken to facilitate vaccine antigen discovery, and to allow for comparative analysis with the related...
Terkawi, Mohamad Alaa; Huyen, Nguyen Xuan; Shinuo, Cao; Inpankaew, Tawin; Maklon, Khuanwalai; Aboulaila, Mahmoud; Ueno, Akio; Goo, Youn-Kyoung; Yokoyama, Naoaki; Jittapalapong, Sathaporn; Xuan, Xuenan; Igarashi, Ikuo
Bovine babesiosis is a tick-transmitted hemoprotozoan disease that is mainly caused by Babesia bovis and Babesia bigemina and is characterized by significant morbidity and mortality worldwide. The disease is widespread in the northeastern region of Thailand, where an increasingly large part of the livestock is composed of water buffaloes. The present study was therefore conducted to investigate the epidemiological distribution of B. bovis and B. bigemina in water buffaloes in the northeastern region of Thailand. A total of 305 buffalo blood samples were randomly collected from five provinces and simultaneously analyzed by the nested PCR (nPCR) assay, ELISA, and IFAT techniques. The overall prevalence of B. bovis and B. bigemina was 11.2% and 3.6% by nPCR, 14.7% and 5.9% by ELISA, and 16.8% and 5.6% by IFAT, respectively. The high concordance between the molecular and the serological detection tests revealed the specificity and sensitivity of the diagnostic assays used for the detection of infection as well as the endemic stability status of the parasites in the surveyed areas. Statistically significant differences in the prevalence of the two infections were observed on the basis of age and location but not gender. Our data provide valuable information regarding the epidemiology of B. bovis and B. bigemina infection in water buffaloes in the northeastern region of Thailand which will likely be very beneficial for management and control programs of this disease.
da Silva, Jenevaldo Barbosa; André, Marcos Rogério; da Fonseca, Adivaldo Henrique; de Albuquerque Lopes, Cinthia Távora; da Silva Lima, Danillo Henrique; de Andrade, Stefano Juliano Tavares; Oliveira, Carlos Magno Chaves; Barbosa, José Diomedes
Bovine babesiosis is a tick-borne disease caused mainly by Babesia bovis and Babesia bigemina, which are associated to considerable economic losses in cattle herds worldwide. Approximately 60% of buffalo herds in South America are located in Northern Brazil. Little is known about the impact of babesiosis on buffalo herds in Brazil. The present work aimed to verify the occurrence of B. bovis and B. bigemina in 542 water buffaloes in the state of Pará, Northern Brazil, using molecular and serological techniques. The percentage of seropositive animals for B. bovis and B. bigemina was 41.2% and 19.0%, respectively, by ELISA. B. bovis and B. bigemina DNA were detected in 15 and 16% of sampled buffaloes, respectively. A high correlation (Kappa index of 0.9) between serological and molecular tests suggests that the combination of the utilized techniques in the present study is suitable for babesiosis diagnosis in an endemic unstable area. Significantly difference of positivity for serological and molecular assays was verified to localities and reproductive status of sampled animals, but not between buffalo breeds. The immune status of sampled buffaloes associated to the circulation of babesiosis agents in sampled population suggests that the studied area is at risk to clinical babesiosis outbreaks. Furthermore, this study demonstrated that this region can be classified as endemically unstable.
Cantu-C, Antonio; Ortega-S, J Alfonso; García-Vázquez, Zeferino; Mosqueda, Juan; Henke, Scott E; George, John E
Species of Rhipicephalus (Boophilus) ticks are the vectors of babesiosis (cattle fever tick), which are distributed worldwide. White-tailed deer (Odocoileus virginianus) are important secondary hosts for the cattle fever ticks, Rhipicephalus (B.) annulatus and Rhipicephalus (B.) microplus. White-tailed deer are capable of sustaining Boophilus spp. tick populations in the presence or absence of cattle. The objectives of this study were to determine the frequency of Babesia bovis and Babesia bigemina and the prevalence of antibodies to them and identify possible risk factors for bovine babesiosis in white-tailed deer in 3 northeastern states of México. Whole blood and serum samples (n = 457) were collected from white-tailed deer in the states of Coahuila, Nuevo Leon, and Tamaulipas during the spring of 2004. Samples were tested for B. bovis and B. bigemina by nested polymerase chain reaction (n-PCR) (the primers for B. bovis identified the gene Rap-1 and B. bigemina were specific primers) and by an indirect immunofluorescence antibody test (IFAT). A questionnaire was given to each ranch to obtain information about management practices. Logistic regression methods were used to test the association between management factors and the dependent variable of positive n-PCR or IFAT. Nineteen (4.2%) samples were positive to B. bigemina and 6 (1.7%) were positive to B. bovis by n-PCR. Serological testing showed 59.9% (n = 274) of deer sampled were positive to B. bovis and 5.4% (n = 25) were positive to B. bigemina antibodies. The logistic model varied with different dependent variables. With positive n-PCR and B. bigemina as the dependent variable, 3 factors were associated: habitat (presence of brush and exotic grasses; odds ratio (OR), 3.3; 95% confidence interval (CI), 1.3-8.5), grazing system (continuous grazing OR 4.0; CI, 1.3-12.2), and tick treatment frequency (3-4 mo; OR 7.0, CI 1.4-34.3; 5-6 mo; OR, 11.0; CI, 1.9-62.7; > 6 mo; OR, 4.6; CI, 0.9-23.3). These findings
Completion of the Babesia bovis (T2Bo strain) genome provides detailed data concerning the predicted proteome of this parasite, and allows for a bioinformatics approach to gene discovery. Comparative genomics of the hemoprotozoan parasites B. bovis and Theileria parva revealed a highly conserved syn...
A novel Babesia bovis gene family encoding proteins with similarities to the Plasmodium 6cys protein family was identified by TBLASTN searches of the Babesia bovis genome using the sequence of the P. falciparum PFS230 protein as query, and was termed Bbo-6cys gene family. The Bbo-cys6 gene family co...
MOSSAAD, Ehab; ASADA, Masahito; NAKATANI, Daichi; INOUE, Noboru; YOKOYAMA, Naoaki; KANEKO, Osamu; KAWAZU, Shin-ichiro
Bovine babesiosis is a livestock disease known to cause economic losses in endemic areas. The apicomplexan parasite Babesia bovis is able to invade and destroy the host’s erythrocytes leading to the serious pathologies of the disease, such as anemia and hemoglobinuria. Understanding the egress mechanisms of this parasite is therefore a key step to develop new therapeutic strategies. In this study, the possible involvement of Ca2+ in the egress of B. bovis merozoites from infected erythrocytes was investigated. Egress was artificially induced in vitro using calcium ionophore A23187 and thapsigargin to increase Ca2+ concentration in the cytosol of the parasite cells. The increased intracellular Ca2+ concentration following these treatments was confirmed using live cell Ca2+ imaging with confocal laser scanning microscopy. Based on our findings, we suggest a Ca2+ signalling pathway in the egress of B. bovis merozoites. PMID:25298241
tBabesia bovis and Babesia bigemina are causative agents of bovine babesiosis, a tick-borne disease of cattlein tropical and subtropical regions. Babesia spp. infection adversely affects cattle health and can be fatalresulting in considerable economic loss worldwide. Under endemic stability conditio...
Munkhjargal, Tserendorj; Ishizaki, Takahiro; Guswanto, Azirwan; Takemae, Hitoshi; Yokoyama, Naoaki; Igarashi, Ikuo
Aminopeptidases are increasingly being investigated as therapeutic targets in various diseases. In this study, we cloned, expressed, and biochemically characterized a member of the methionine aminopeptidase (MAP) family from Babesia bovis (B. bovis) to develop a potential molecular drug target. Recombinant B. bovis MAP (rBvMAP) was expressed in Escherichia coli (E. coli) as a glutathione S-transferase (GST)-fusion protein, and we found that it was antigenic. An antiserum against the rBvMAP protein was generated in mice, and then a native B. bovis MAP was identified in B. bovis by Western blot assay. Further, an immunolocalization assay showed that MAP is present in the cytoplasm of the B. bovis merozoite. Analysis of the biochemical properties of rBvMAP revealed that it was enzymatically active, with optimum activity at pH 7.5. Enhanced enzymatic activity was observed in the presence of divalent manganese cations and was effectively inhibited by a metal chelator, ethylenediaminetetraacetic acid (EDTA). Moreover, the enzymatic activity of BvMAP was inhibited by amastatin and bestatin as inhibitors of MAP (MAPi) in a dose-dependent manner. Importantly, MAPi was also found to significantly inhibit the growth of Babesia parasites both in vitro and in vivo; additionally, they induced high levels of cytokines and immunoglobulin (IgG) titers in the host. Therefore, our results suggest that BvMAP is a molecular target of amastatin and bestatin, and those inhibitors may be drug candidates for the treatment of babesiosis, though more studies are required to confirm this.
Incidence of bovine babesiosis in Portugal is currently unknown. In this study, a first survey of Babesia bovis and B. bigemina infection in cattle was carried out using blood samples from 406 clinically healthy individuals from different districts from Central and Southern regions of Portugal and a...
Genis, Alma D; Mosqueda, Juan J; Borgonio, Verónica M; Falcón, Alfonso; Alvarez, Antonio; Camacho, Minerva; de Lourdes Muñoz, Maria; Figueroa, Julio V
Variable merozoite surface antigens of Babesia bovis are exposed glycoproteins having a role in erythrocyte invasion. Members of this gene family include msa-1 and msa-2 (msa-2c, msa-2a(1), msa-2a(2), and msa-2b). Small subunit ribosomal (ssr)RNA gene is subject to evolutive pressure and has been used in phylogenetic studies. To determine the phylogenetic relationship among B. bovis Mexican isolates using different genetic markers, PCR amplicons, corresponding to msa-1, msa-2c, msa-2b, and ssrRNA genes, were cloned and plasmids carrying the corresponding inserts were sequenced. Comparative analysis of nucleotide and deduced amino acid sequences revealed distinct degrees of variability and identity among the coding gene sequences obtained from 12 geographically different B. bovis isolates and a reference strain. Overall sequence identities of 47.7%, 72.3%, 87.7%, and 94% were determined for msa-1, msa-2b, msa-2c, and ssrRNA, respectively. A robust phylogenetic tree was obtained with msa-2b sequences. The phylogenetic analysis suggests that Mexican B. bovis isolates group in clades not concordant with the Mexican geography. However, the Mexican isolates group together in an American clade separated from the Australian clade. Sequence heterogeneity in msa-1, msa-2b, and msa-2c coding regions of Mexican B. bovis isolates present in different geographical regions can be a result of either differential evolutive pressure or cattle movement from commercial trade.
Begley, Darren W.; Edwards, Thomas E.; Raymond, Amy C.; Smith, Eric R.; Hartley, Robert C.; Abendroth, Jan; Sankaran, Banumathi; Lorimer, Donald D.; Myler, Peter J.; Staker, Bart L.; Stewart, Lance J.
Babesiosis is a tick-borne disease caused by eukaryotic Babesia parasites which are morphologically similar to Plasmodium falciparum, the causative agent of malaria in humans. Like Plasmodium, different species of Babesia are tuned to infect different mammalian hosts, including rats, dogs, horses and cattle. Most species of Plasmodium and Babesia possess an essential bifunctional enzyme for nucleotide synthesis and folate metabolism: dihydrofolate reductase-thymidylate synthase. Although thymidylate synthase is highly conserved across organisms, the bifunctional form of this enzyme is relatively uncommon in nature. The structural characterization of dihydrofolate reductase-thymidylate synthase in Babesia bovis, the causative agent of babesiosis in livestock cattle, is reported here. The apo state is compared with structures that contain dUMP, NADP and two different antifolate inhibitors: pemetrexed and raltitrexed. The complexes reveal modes of binding similar to that seen in drug-resistant malaria strains and point to the utility of applying structural studies with proven cancer chemotherapies towards infectious disease research. PMID:21904052
Asada, Masahito; Goto, Yasuyuki; Yahata, Kazuhide; Yokoyama, Naoaki; Kawai, Satoru; Inoue, Noboru; Kaneko, Osamu; Kawazu, Shin-ichiro
Background Babesia bovis is an apicomplexan intraerythrocytic protozoan parasite that induces babesiosis in cattle after transmission by ticks. During specific stages of the apicomplexan parasite lifecycle, such as the sporozoites of Plasmodium falciparum and tachyzoites of Toxoplasma gondii, host cells are targeted for invasion using a unique, active process termed “gliding motility”. However, it is not thoroughly understood how the merozoites of B. bovis target and invade host red blood cells (RBCs), and gliding motility has so far not been observed in the parasite. Methodology/Principal Findings Gliding motility of B. bovis merozoites was revealed by time-lapse video microscopy. The recorded images revealed that the process included egress of the merozoites from the infected RBC, gliding motility, and subsequent invasion into new RBCs. The gliding motility of B. bovis merozoites was similar to the helical gliding of Toxoplasma tachyzoites. The trails left by the merozoites were detected by indirect immunofluorescence assay using antiserum against B. bovis merozoite surface antigen 1. Inhibition of gliding motility by actin filament polymerization or depolymerization indicated that the gliding motility was driven by actomyosin dependent process. In addition, we revealed the timing of breakdown of the parasitophorous vacuole. Time-lapse image analysis of membrane-stained bovine RBCs showed formation and breakdown of the parasitophorous vacuole within ten minutes of invasion. Conclusions/Significance This is the first report of the gliding motility of B. bovis. Since merozoites of Plasmodium parasites do not glide on a substrate, the gliding motility of B. bovis merozoites is a notable finding. PMID:22506073
Oldiges, Daiane P.; Laughery, Jacob M.; Tagliari, Nelson Junior; Leite Filho, Ronaldo Viana; Davis, William C.; da Silva Vaz, Itabajara; Termignoni, Carlos; Knowles, Donald P.; Suarez, Carlos E.
The Rhipicephalus microplus tick is a notorious blood-feeding ectoparasite of livestock, especially cattle, responsible for massive losses in animal production. It is the main vector for transmission of pathogenic bacteria and parasites, including Babesia bovis, an intraerythrocytic apicomplexan protozoan parasite responsible for bovine Babesiosis. This study describes the development and testing of a live B. bovis vaccine expressing the protective tick antigen glutathione-S-transferase from Haemaphysalis longicornis (HlGST). The B. bovis S74-T3B parasites were electroporated with a plasmid containing the bidirectional Ef-1α (elongation factor 1 alpha) promoter of B. bovis controlling expression of two independent genes, the selectable marker GFP-BSD (green fluorescent protein–blasticidin deaminase), and HlGST fused to the MSA-1 (merozoite surface antigen 1) signal peptide from B. bovis. Electroporation followed by blasticidin selection resulted in the emergence of a mixed B. bovis transfected line (termed HlGST) in in vitro cultures, containing parasites with distinct patterns of insertion of both exogenous genes, either in or outside the Ef-1α locus. A B. bovis clonal line termed HlGST-Cln expressing intracellular GFP and HlGST in the surface of merozoites was then derived from the mixed parasite line HlGST using a fluorescent activated cell sorter. Two independent calf immunization trials were performed via intravenous inoculation of the HlGST-Cln and a previously described control consisting of an irrelevant transfected clonal line of B. bovis designated GFP-Cln. The control GFP-Cln line contains a copy of the GFP-BSD gene inserted into the Ef-1α locus of B. bovis in an identical fashion as the HIGST-Cln parasites. All animals inoculated with the HlGST-Cln and GFP-Cln transfected parasites developed mild babesiosis. Tick egg fertility and fully engorged female tick weight was reduced significantly in R. microplus feeding on HlGST-Cln-immunized calves
Tattiyapong, Muncharee; Sivakumar, Thillaiampalam; Takemae, Hitoshi; Simking, Pacharathon; Jittapalapong, Sathaporn; Igarashi, Ikuo; Yokoyama, Naoaki
Babesia bovis, an intraerythrocytic protozoan parasite, causes severe clinical disease in cattle worldwide. The genetic diversity of parasite antigens often results in different immune profiles in infected animals, hindering efforts to develop immune control methodologies against the B. bovis infection. In this study, we analyzed the genetic diversity of the merozoite surface antigen-1 (msa-1) gene using 162 B. bovis-positive blood DNA samples sourced from cattle populations reared in different geographical regions of Thailand. The identity scores shared among 93 msa-1 gene sequences isolated by PCR amplification were 43.5-100%, and the similarity values among the translated amino acid sequences were 42.8-100%. Of 23 total clades detected in our phylogenetic analysis, Thai msa-1 gene sequences occurred in 18 clades; seven among them were composed of sequences exclusively from Thailand. To investigate differential antigenicity of isolated MSA-1 proteins, we expressed and purified eight recombinant MSA-1 (rMSA-1) proteins, including an rMSA-1 from B. bovis Texas (T2Bo) strain and seven rMSA-1 proteins based on the Thai msa-1 sequences. When these antigens were analyzed in a western blot assay, anti-T2Bo cattle serum strongly reacted with the rMSA-1 from T2Bo, as well as with three other rMSA-1 proteins that shared 54.9-68.4% sequence similarity with T2Bo MSA-1. In contrast, no or weak reactivity was observed for the remaining rMSA-1 proteins, which shared low sequence similarity (35.0-39.7%) with T2Bo MSA-1. While demonstrating the high genetic diversity of the B. bovis msa-1 gene in Thailand, the present findings suggest that the genetic diversity results in antigenicity variations among the MSA-1 antigens of B. bovis in Thailand.
Objective: The Babesia bovis genome encodes a rap-1 related gene denominated RAP-1 related antigen (RRA). In this study, we analyzed the pattern of expression, immunogenicity and functional relevance of RRA. Methods: Phylogenetic analysis was performed using the program Phylip. Expression of rra wa...
Small open reading frame (smorf) genes comprise the second largest Babesia bovis multigene family. All known 44 variant smorf genes are located in close chromosomal proximity to ves1 genes, which encode proteins that mediate cytoadhesion and contribute to immune evasion. In this study, we characte...
Babesia bovis caused bovine babesiosis is a world tick borne hemoprotozoan disease leading to fever, anemia, weight losses and ultimately death. Several babesicidal drugs that have been in use in cattle for years have proven to be partially ineffective and the development of alternative highly speci...
Genis, Alma D; Perez, Jocelin; Mosqueda, Juan J; Alvarez, Antonio; Camacho, Minerva; Muñoz, Maria de Lourdes; Rojas, Carmen; Figueroa, Julio V
Variable merozoite surface antigens of Babesia bovis are exposed glycoproteins having a role in erythrocyte invasion. Members of this gene family include msa-1 and msa-2 (msa-2c, msa-2a(1), msa-2a(2) and msa-2b). To determine the sequence variation among B. bovis Mexican isolates using msa-2b as a genetic marker, PCR amplicons corresponding to msa-2b were cloned and plasmids carrying the corresponding inserts were purified and sequenced. Comparative analysis of nucleotide and deduced amino acid sequences revealed distinct degrees of variability and identity among the coding gene sequences obtained from 16 geographically different Mexican B. bovis isolates and a reference strain. Clustal-W multiple alignments of the MSA-2b deduced amino acid sequences performed with the 17 B. bovis Mexican isolates, revealed the identification of three genotypes with a distinct set each of amino acid residues present at the variable region: Genotype I represented by the MO7 strain (in vitro culture-derived from the Mexico isolate) as well as RAD, Chiapas-1, Tabasco and Veracruz-3 isolates; Genotype II, represented by the Jalisco, Mexico and Veracruz-2 isolates; and Genotype III comprising the sequences from most of the isolates studied, Tamaulipas-1, Chiapas-2, Guerrero-1, Nayarit, Quintana Roo, Nuevo Leon, Tamaulipas-2, Yucatan and Guerrero-2. Moreover, these three genotypes could be discriminated against each other by using a PCR-RFLP approach. The results suggest that occurrence of indels within the variable region of msa-2b sequences can be useful markers for identifying a particular genotype present in field populations of B. bovis isolated from infected cattle in Mexico.
Rojas-Martínez, C; Rodríguez-Vivas, R I; Figueroa Millán, J V; Acosta Viana, K Y; Gutiérrez Ruiz, E J; Álvarez Martínez, J A
This study reports the effect of putrescine addition, either alone or in combination with insulin, transferrin and selenite (ITS), to serum-free Advanced DMEM/F12 (A-DMEM/F12) medium, on the in vitro culture of Babesia bovis and using a perfusion bioreactor to improve efficiency of the process. A B. bovis strain previously adapted to proliferate in serum-free medium (Bbovis-SF) was evaluated using eight increasing concentrations of putrescine. The percentage of parasitized erythrocytes (PPE) obtained from cultures supplemented with 0.101 mg/L was 6.23% compared with 2.3% for control cultures with M199 with Earle's salts (M199) and 40% serum. The combination of putrescine (0.101 mg/L) and a mixture of ITS (2000, 1100, and 1.34 mg/L, respectively) (Pu-ITS), in A-DMEM/F12 culture medium without serum yielded a maximum PPE of 17.26% compared to 2.58% in the control medium. This new formulation of culture medium, together with the use of a hollow-fiber perfusion bioreactor system (HFPBS), caused a substantial increase in the proliferation of B. bovis, yielding a maximum cumulative PPE of 118.8% after five days, compared to 58.6% in cultures treated with control medium M199 and 40% serum. We concluded that the addition of the ITS mixture and putrescine to the culture medium stimulated the proliferation of B. bovis in vitro. This new medium formulation, used in a HFPBS culture system, can be an effective, automated-prone system that can induce massive proliferation of B. bovis for use as a source of parasite antigens and immunogens.
Ibrahim, Hany M; Adjou Moumouni, Paul F; Mohammed-Geba, Khaled; Sheir, Sherin K; Hashem, Ihab S Y; Cao, Shinuo; Terkawi, Mohamad A; Kamyingkird, Ketsarin; Nishikawa, Yoshifumi; Suzuki, Hiroshi; Xuan, Xuenan
In order to determine the molecular and serological prevalence of Babesia bigemina and Babesia bovis, a total of 247 blood samples were collected from cattle and water buffalos in Beheira and Faiyum Provinces in Egypt and examined by nested polymerase chain reaction (nPCR) and enzyme-linked immunosorbent assay (ELISA). In cattle, the prevalence of B. bigemina and B. bovis was 5.30% and 3.97% by nPCR and 10.60% and 9.27% by ELISA, respectively, whereas those of water buffalos were 10.42% and 4.17% by nPCR and 15.63% and 11.46% by ELISA, respectively. Statistically significant differences in the prevalence of the two infections were observed on the basis of age and health status. Sequencing analysis revealed two genotypes for B. bovis spherical body protein-4. In conclusion, the current data provide valuable information regarding the epidemiology of B. bigemina and B. bovis infections in cattle and water buffalos from Egypt, which can be employed in developing future strategies for disease management and control.
Romero-Salas, Dora; Mira, Anabela; Mosqueda, Juan; García-Vázquez, Zeferino; Hidalgo-Ruiz, Mario; Vela, Noot Aditya Ortiz; de León, Adalberto Angel Perez; Florin-Christensen, Monica; Schnittger, Leonhard
Babesia bovis and Babesia bigemina are causative agents of bovine babesiosis, a tick-borne disease of cattle in tropical and subtropical regions. Babesia spp. infection adversely affects cattle health and can be fatal resulting in considerable economic loss worldwide. Under endemic stability conditions, herds contain high numbers of chronically infected, asymptomatic carrier animals, in which no parasitemia is detected by microscopic blood smear examination. In addition to bovines, also water buffaloes are infected by both Babesia spp. commonly leading to a subclinical infection. The infection rate (by nPCR) and herd exposure (by IFAT) of bovines and water buffaloes reared under similar field conditions in an area of endemic stability were determined and compared. In order to optimize direct parasite detection, highly sensitive nPCR assays were developed and applied, allowing the detection of as little as 0.1 fg DNA of each Babesia pathogen. Significantly lower percentages (p<0.001) of seropositive water buffaloes compared to bovines were observed for B. bovis (71.4% vs. 98%) and B. bigemina (85% vs. 100%). Interestingly, in comparison, differences noticed between water buffaloes and bovines were considerably larger with direct parasite detection by nPCR (16.2% vs. 82.3% and 24% vs. 94.1% for B. bovis and B. bigemina, respectively). As expected, bovines subjected to monthly acaricide applications exhibited a significant lower infection rate as determined by nPCR than bovines not subjected to these measures (B. bovis 33.3% vs. 90.7%, p<0.001; B. bigemina 80% vs. 96.5%, p<0.001, for treated vs. untreated animals). Interestingly no differences between these groups were observed with respect to seropositivity, suggesting similar rates of parasite exposure (B. bovis 100% vs. 97.7%, p<0.001; B. bigemina 100% vs. 100%, p<0.001). Importantly, a significantly higher number of water buffaloes as determined by nPCR were infected when reared jointly with bovines not subjected
Genetic manipulation is an essential technique to analyze gene function; however, limited methods are available for Babesia bovis, a causative pathogen of the globally important cattle disease, bovine babesiosis. To date, two stable transfection systems have been developed for B. bovis, using select...
Background Cattle babesiosis is a tick-borne disease of cattle that has severe economic impact on cattle producers throughout the world’s tropical and subtropical countries. The most severe form of the disease is caused by the apicomplexan, Babesia bovis, and transmitted to cattle through the bite of infected cattle ticks of the genus Rhipicephalus, with the most prevalent species being Rhipicephalus (Boophilus) microplus. We studied the reaction of the R. microplus larval transcriptome in response to infection by B. bovis. Methods Total RNA was isolated for both uninfected and Babesia bovis-infected larval samples. Subtracted libraries were prepared by subtracting the B. bovis-infected material with the uninfected material, thus enriching for expressed genes in the B. bovis-infected sample. Expressed sequence tags from the subtracted library were generated, assembled, and sequenced. To complement the subtracted library method, differential transcript expression between samples was also measured using custom high-density microarrays. The microarray probes were fabricated using oligonucleotides derived from the Bmi Gene Index database (Version 2). Array results were verified for three target genes by real-time PCR. Results Ticks were allowed to feed on a B. bovis-infected splenectomized calf and on an uninfected control calf. RNA was purified in duplicate from whole larvae and subtracted cDNA libraries were synthesized from Babesia-infected larval RNA, subtracting with the corresponding uninfected larval RNA. One thousand ESTs were sequenced from the larval library and the transcripts were annotated. We used a R. microplus microarray designed from a R. microplus gene index, BmiGI Version 2, to look for changes in gene expression that were associated with infection of R. microplus larvae. We found 24 transcripts were expressed at a statistically significant higher level in ticks feeding upon a B. bovis-infected calf contrasted to ticks feeding on an uninfected calf
Combrink, M P; Troskie, P C; Pienaar, R; Latif, A A; Mans, B J
Genotypic diversity in Babesia bovis (cause of Asiatic redwater in cattle) vaccine strains and field isolates from South Africa were investigated using the Bv80 gene as well as microsatellites. The S11 vaccine strain possessed both A and B alleles of the Bv80 gene, as well as genotypic diversity within each allele type as defined by repeat variation resulting in different amplicon sizes. Rapid serial passage of vaccine strain from passage S10 to S24 resulted in loss of genotypic diversity that yielded a single allele A genotype with an amplicon size of 558 bp. This suggested that clonal selection occurred during rapid passaging. Extensive genotypic diversity exists in 44 field isolates characterized with both Bv80 A and B alleles, but can be readily distinguished from the S24 vaccine strain using either the Bv80 allele specific PCR assays or using multi-locus micro-satellite typing. This indicated that no recent documented clinical cases of Asiatic redwater were caused by the reversion to virulence of the current vaccine strain.
Simking, Pacharathon; Saengow, Sinsamuth; Bangphoomi, Kunan; Sarataphan, Nachai; Wongnarkpet, Sirichai; Inpankaew, Tawin; Jittapalapong, Sathaporn; Munkhjargal, Tserendorj; Sivakumar, Thillaiampalam; Yokoyama, Naoaki; Igarashi, Ikuo
Bovine babesiosis is an economically significant disease that affects dairy farming operations in Thailand. In the present study, 1824 blood-DNA samples prepared from cattle bred in 4 different regions of the country (North, Northeast, Central, and South) were screened using a nested PCR for the specific detection of Babesia bovis. While the overall prevalence of B. bovis was 8.8%, the Central region of Thailand was found to be a high-risk area of the country, as the prevalence of the parasite was 15.0%. The positive rate was relatively higher among the animals of 1-5 years of age. The genetic diversity among the B. bovis parasites was also studied based on their MSA-2b gene, and the findings showed that the Thai sequences were dispersed across 8 of 13 total clades observed in the phylogram. Three of these clades were formed only of Thai sequences. Similarity among the deduced MSA-2b amino acid sequences determined in the present study was 68.3-100%. In conclusion, the present study found that all the locations surveyed were infected with B. bovis and that the parasite populations in Thailand were genetically diverse. Our findings highlight the need for further studies in Thailand to generate more information before a sound control strategy could be implemented against B. bovis.
Harrus, S; Perlman-Avrahami, A; Mumcuoglu, K Y; Morick, D; Eyal, O; Baneth, G
: Ticks are vectors of important pathogens of human and animals. Therefore, their microbial carriage capacity is constantly being investigated. The aim of this study was to characterize the diversity of domestic animal pathogens in ticks collected from vegetation and the ground, from different parts of Israel. Non-engorged questing adult ticks were collected from 13 localities. A total of 1196 ticks in 131 pools-83 pools of Rhipicephalus turanicus and 48 of Rhipicephalus sanguineus (with two to ten ticks per pool)-were included in this study. In addition, 13 single free-roaming Hyalomma spp. ticks were collected. Screening by molecular techniques revealed the presence of Ehrlichia canis, Anaplasma platys, Anaplasma bovis and Babesia canis vogeli DNA in R. turanicus ticks. E. canis, A. bovis, B. canis vogeli and Candidatus Midichloria mitochondrii DNA sequences were detected in R. sanguineus ticks. Candidatus Midichloria mitochondrii DNA was also detected in Hyalomma spp. ticks. Neither Hepatozoon spp. nor Bartonella spp. DNA was detected in any of the ticks examined. This study describes the first detection of E. canis in the tick R. turanicus, which may serve as a vector of this canine pathogen; E. canis was the most common pathogen detected in the collected questing ticks. It also describes the first detection of A. bovis and Candidatus Midichloria mitochondrii in Israel. To the best of the author's knowledge, this is the first report describing the detection of DNA of the latter two pathogens in R. sanguineus, and of A. bovis in R. turanicus.
Background: Tick borne Babesia bigemina is responsible for acute and potentially lethal hemolytic disease in cattle. The development of genetic manipulation tools necessary to the better understanding of parasite biology is currently limited by the lack of a complete parasite genome and experimental...
Boophilus ticks are vectors of Babesia bovis, the protozoan causative agent of cattle fever, a disease which is responsible for significant production losses to cattle producers in much of Africa, Central and South America and Australia. We utilized subtractive cDNA library synthesis techniques to o...
Boophilus ticks are vectors of Babesia bovis, the protozoan causative agent of cattle fever, a disease which is responsible for significant production losses to cattle producers in much of Africa, Central and South America and Australia. We utilized R. microplus microarrays, which contained over 13,...
Background Cattle babesiosis is a tick-borne disease of cattle with the most severe form of the disease caused by the apicomplexan, Babesia bovis. Babesiosis is transmitted to cattle through the bite of infected cattle ticks of the genus Rhipicephalus. The most prevalent species is Rhipicephalus (Boophilus) microplus, which is distributed throughout the tropical and subtropical countries of the world. The transmission of B. bovis is transovarian and a previous study of the R. microplus ovarian proteome identified several R. microplus proteins that were differentially expressed in response to infection. Through various approaches, we studied the reaction of the R. microplus ovarian transcriptome in response to infection by B. bovis. Methods A group of ticks were allowed to feed on a B. bovis-infected splenectomized calf while a second group fed on an uninfected splenectomized control calf. RNA was purified from dissected adult female ovaries of both infected and uninfected ticks and a subtracted B. bovis-infected cDNA library was synthesized, subtracting with the uninfected ovarian RNA. Four thousand ESTs were sequenced from the ovary subtracted library and annotated. Results The subtracted library dataset assembled into 727 unique contigs and 2,161 singletons for a total of 2,888 unigenes, Microarray experiments designed to detect B. bovis-induced gene expression changes indicated at least 15 transcripts were expressed at a higher level in ovaries from ticks feeding upon the B. bovis-infected calf as compared with ovaries from ticks feeding on an uninfected calf. We did not detect any transcripts from these microarray experiments that were expressed at a lower level in the infected ovaries compared with the uninfected ovaries. Using the technique called serial analysis of gene expression, 41 ovarian transcripts from infected ticks were differentially expressed when compared with transcripts of controls. Conclusion Collectively, our experimental approaches provide
Borgonio, Veronica; Mosqueda, Juan; Genis, Alma D; Falcon, Alfonso; Alvarez, J Antonio; Camacho, Minerva; Figueroa, Julio V
Babesia bovis msa-1 and msa-2c genes belong to the variable merozoite surface antigen gene family. These genes code for antigenic proteins present on the merozoite surface (MSA) and are involved in the parasite invasion to the bovine erythrocyte. Previous studies carried out on MSA-1 have evidenced antigen allelic variation in B. bovis isolates from similar endemic regions, as well as in isolates from different geographic regions of the world (Argentina, Australia, Israel). Studies conducted on MSA-2c, however, have shown that this antigen is widely conserved on isolates from distinct geographic regions. In this study, it was hypothesized that MSA-1 and MSA-2c antigens would contain common epitopes despite the presence of nucleotide sequence differences found in 13 B. bovis isolates and strains collected in geographically distant regions of Mexico. Bioinformatics analysis of the primary structure from DNA fragments derived from PCR amplification, cloning, and sequencing of msa-1 and msa-2c genes from the 13 B. bovis populations revealed that the msa-1 gene product present in the various isolates tested is less conserved among isolates obtained within a similar geographic region in Mexico (51-99.7% sequence identity). Results obtained by immunoblot analysis of B. bovis protein extracts reacted with a monoclonal antibody to MSA-1 42-kDa antigen, conclusively showed cross-reactive common epitopes only in Mexican isolates having high sequence identity (>/=99%, eight isolates). Sequence analysis and multiple alignment of deduced MSA-2c demonstrated a high degree of sequence identity (90-100%) among the Mexican B. bovis isolates and strains. Immunoblot results using a polyclonal antibody to MSA-2c reacted against the protein extracts recognized conserved epitopes in at least nine of the B. bovis isolates. The results obtained in this study agree with those previously reported by other researchers and confirm that, based in sequence identity conservation in Mexican B
Yokoyama, Naoaki; Sivakumar, Thillaiampalam; Tuvshintulga, Bumduuren; Hayashida, Kyoko; Igarashi, Ikuo; Inoue, Noboru; Long, Phung Thang; Lan, Dinh Thi Bich
The genes that encode merozoite surface antigens (MSAs) in Babesia bovis are genetically diverse. In this study, we analyzed the genetic diversity of B. bovis MSA-1, MSA-2b, and MSA-2c genes in Vietnamese cattle and water buffaloes. Blood DNA samples from 258 cattle and 49 water buffaloes reared in the Thua Thien Hue province of Vietnam were screened with a B. bovis-specific diagnostic PCR assay. The B. bovis-positive DNA samples (23 cattle and 16 water buffaloes) were then subjected to PCR assays to amplify the MSA-1, MSA-2b, and MSA-2c genes. Sequencing analyses showed that the Vietnamese MSA-1 and MSA-2b sequences are genetically diverse, whereas MSA-2c is relatively conserved. The nucleotide identity values for these MSA gene sequences were similar in the cattle and water buffaloes. Consistent with the sequencing data, the Vietnamese MSA-1 and MSA-2b sequences were dispersed across several clades in the corresponding phylogenetic trees, whereas the MSA-2c sequences occurred in a single clade. Cattle- and water-buffalo-derived sequences also often clustered together on the phylogenetic trees. The Vietnamese MSA-1, MSA-2b, and MSA-2c sequences were then screened for recombination with automated methods. Of the seven recombination events detected, five and two were associated with the MSA-2b and MSA-2c recombinant sequences, respectively, whereas no MSA-1 recombinants were detected among the sequences analyzed. Recombination between the sequences derived from cattle and water buffaloes was very common, and the resultant recombinant sequences were found in both host animals. These data indicate that the genetic diversity of the MSA sequences does not differ between cattle and water buffaloes in Vietnam. They also suggest that recombination between the B. bovis MSA sequences in both cattle and water buffaloes might contribute to the genetic variation in these genes in Vietnam.
Suarez, Carlos E; Johnson, Wendell C; Herndon, David R; Laughery, Jacob M; Davis, William C
This study examines the patterns of gene integration of gfp-bsd upon stable transfection into the T3Bo strain of Babesia bovis using a plasmid designed to integrate homologous sequences of the parasite's two identical ef-1α A and B genes. While the transfected BboTf-149-6 cell line displayed two distinct patterns of gene integration, clonal lines derived from this strain by cell sorting contained only single gfp-bsd insertions. Whole genome sequencing of two selected clonal lines, E9 and C6, indicated two distinct patterns of gfp-bsd insertion occurring by legitimate homologous recombination mechanisms: one into the expected ef-1α orf B, and another into the ef-1α B promoter. The data suggest that expression of the ef-1α orf B is not required for development of B. bovis in cultured erythrocyte stages. Use of legitimate homologous recombination mechanisms in transfected B. bovis supports the future use of transfection methods for developing efficient gene function assignment experiments using gene knockout techniques.
Dominguez, Mariana; Echaide, Ignacio; de Echaide, Susana Torioni; Wilkowsky, Silvina; Zabal, Osvaldo; Mosqueda, Juan J.; Schnittger, Leonhard
Infections by Babesia bovis limit cattle production and cause important economic losses in tropical and subtropical areas around the world. Monitoring of calf sera can be used to detect unprotected cattle herds and to decide on strategic control measures, as well as for epidemiological studies. Merozoite surface antigen 2c (MSA-2c) is an immunodominant surface protein expressed in B. bovis merozoites and sporozoites and contains B-cell epitopes that are conserved among geographic isolates. A monoclonal antibody against recombinant MSA-2c (rMSA-2c) was previously shown to inhibit the binding of anti-B. bovis antibodies to a parasite B-cell epitope in a competitive enzyme-linked immunosorbent assay (cELISA) format. In the work at hand, the parameters of this cELISA were reevaluated and adjusted when necessary, and a cutoff value was determined by receiver operator characteristic (ROC) curve analysis of a total of 357 bovine sera of known reactivity, as assessed by indirect immunofluorescence assay (IFAT). The established rMSA-2c cELISA demonstrated a specificity of 98% and a sensitivity of 96.2%. An additional set of 303 field bovine sera from regions where ticks are endemic and tick-free regions of Argentina was tested by both rMSA-2c cELISA and IFAT, and the results were shown to be in very good agreement (kappa index, 0.8325). The performance shown by rMSA-2c cELISA in the detection of B. bovis-specific antibodies and its suitability for standardization and large-scale production, as well as the possibility of its application in most veterinary diagnostic laboratories, make the assay a powerful tool for the surveillance of herd immunity as a strategic measure for the control of bovine babesiosis. PMID:22492742
Liyanagunawardena, Nilukshi; Sivakumar, Thillaiampalam; Kothalawala, Hemal; Silva, Seekkuge Susil Priyantha; Battsetseg, Badgar; Lan, Dinh Thi Bich; Inoue, Noboru; Igarashi, Ikuo; Yokoyama, Naoaki
Babesia bovis is the most virulent Babesia organism, resulting in a high mortality rate in cattle. The genetic diversity of B. bovis merozoite surface antigens (MSAs), such as MSA-1, MSA-2b, and MSA-2c, might be linked to altered immune profiles in the host animals. The present study aimed to develop type-specific PCR assays for Asian msa-1 genotypes, thereby re-analyzing the genetic diversity of msa-1 in Sri Lanka, Mongolia, and Vietnam. Specific primers were designed for nine Asian msa-1 genotypes, which had been detected based on the phylogeny constructed using msa-1 gene sequences retrieved from the GenBank database. Specificity of the type-specific PCR assays was confirmed using plasmids containing the inserts of msa-1 gene fragments that represent Asian genotypes. Furthermore, no amplicons were observed by these PCR assays when DNA samples of Babesia bigemina, Babesia ovata, Theileria annulata, Theileria orientalis, Trypanosoma evansi, Trypanosoma theileri, Anaplasma marginale, and Anaplasma bovis, and non-infected bovine blood were analyzed. In total, 109 B. bovis-positive blood DNA samples sourced from Sri Lanka (44 cattle), Mongolia (26 cattle), and Vietnam (23 cattle and 16 water buffaloes) were then screened by the type-specific PCR assays. The sequences derived from all of the PCR amplicons were phylogenetically analyzed. Out of 109 DNA samples, 23 (20 from cattle and 3 from water buffaloes) were positive for at least one genotype. In agreement with previous studies, five and four different genotypes were detected among the DNA samples from Sri Lanka and Vietnam, respectively. In contrast, four genotypes, including three novel genotypes, were detected from Mongolia. Five DNA samples were found to be co-infected with multiple genotypes. The sequences of the PCR amplicons clustered phylogenetically within the corresponding clades. These findings indicated that the type-specific PCR assays described herein are useful for the determination of genotypic
Rodriguez, Anabel E; Zamorano, Patricia; Wilkowsky, Silvina; Torrá, Florencia; Ferreri, Lucas; Dominguez, Mariana; Florin-Christensen, Mónica
Liposomes prepared from total egg yolk lipid extracts were used to deliver experimental DNA vaccines to mice consisting of pCI-neo plasmids encoding bovine herpesvirus type 1 (BoHV-1) gD or Babesia bovis MSA-2c. A significantly higher proportion of mice in the B. bovis MSA-2c group, but not those in the BoHV-1 gD group, developed detectable immunoglobulin G responses when vaccinated with liposome encapsulated DNA in comparison with mice vaccinated with naked DNA. In both groups, antibody titres were similar between mice vaccinated with liposome encapsulated DNA and naked DNA.
Gohil, Sejal; Kats, Lev M; Seemann, Torsten; Fernandez, Kate M; Siddiqui, Ghizal; Cooke, Brian M
Babesia bovis is a pathogen of considerable economic significance to the livestock industry worldwide but the precise mechanisms by which this parasite causes disease in susceptible cattle remain poorly understood. It is clear, however, that alterations to the structure and function of red blood cells in which the parasites reside and replicate play an important role in pathogenesis and that these are secondary to the export of numerous, currently unknown and uncharacterised parasite-encoded proteins. Using a rational bioinformatic approach, we have identified a set of 362 proteins (117 of which are hypothetical) that we predict encompasses the B. bovis exportome. These exported proteins are likely to be trafficked to various cellular locations, with a subset destined for the red blood cell cytosol or the red blood cell cytoskeleton. These proteins are likely to play important roles in mediating the pathogenesis of babesiosis. We have selected three novel proteins and confirmed their predicted export and localisation within the host red blood cell by immunofluorescence using specific antibodies raised against these proteins. Complete characterisation of these novel exported parasite proteins will help elucidate their function within the host red blood cell and assist in identification of new therapeutic targets for babesiosis.
Antonio Alvarez, J; Lopez, U; Rojas, C; Borgonio, V M; Sanchez, V; Castañeda, R; Vargas, P; Figueroa, J V
The purpose of this research was to evaluate the recombinant proteins MSA-1, MSA-2c and 12D3 as a combined immunogen for cattle. Fifteen steers were randomly assigned into three groups of five animals each (I, II and III). On day 0, cattle in group I were injected with 50 microg each of rMSA-1, rMSA-2c and r12D3 with the adjuvant Montanide 75; cattle in Group II received adjuvant-PBS, and Group III were untreated controls. On day 14, cattle in Group I received a second injection of the three recombinant proteins in adjuvant and cattle in Group II again received adjuvant alone. On day 28, all groups of cattle were challenged with a field strain of Babesia bovis. After challenge, the experimental cattle were clinically and serologically monitored. Three of the five steers immunized with the combined recombinant B. bovis proteins seroconverted on day 14 post-immunization (P.I.) and the maximum titre was 1 : 1600. All five immunized steers presented strong seropositivity to B. bovis antigens at day 21 P.I. The prepatent periods of vaccinated cattle were delayed until day 10 post-challenge exposure versus 8 and 7 days in Groups II and III, respectively. Cattle in all groups had fever above 41 degrees C; the reduction in packed cell volume was not significantly different (P > 0.05) in vaccinated group I compared with Groups II and III (29% versus 26% and 31%, respectively). Treatment was required for one steer in the control group. During the period of the study, the weight of cattle in Groups I and II increased an average of 9 and 7 kg, whereas the weight of the control cattle was reduced on average 4 kg. Immunization with rMSA-1-rMSA-2c-r12D3 proteins was not sufficient to prevent clinical symptoms against challenge, but the immunologic response was sufficient to protect steers against a mild virulent strain of B. bovis.
Rojas Martínez, C; Rodríguez-Vivas, R I; Figueroa Millán, J V; Acosta Viana, K Y; Gutiérrez Ruiz, E J; Álvarez Martínez, J A
Bovine serum is an important factor for the optimal growth of Babesia bovis in vitro. This protozoan can be cultured in M-199 with Earle's salts medium (M-199) supplemented with 40% bovine serum (BS). In the present study, four media were assessed along with the control medium M-199. The effect on the proliferation of B. bovis in vitro was tested when these media were combined with insulin (Ins), transferrin (Trans) and selenite (Sel) in the absence of bovine serum. Treatment with Advanced DMEM/F12 medium (A-DMEM/F12) achieved the highest percentage of parasitized erythrocytes (PPE), reaching a maximum value of 9.59%. A-DMEM/F12 medium supplemented with a mixture of Ins (2000 mg/L), Trans (1100 mg/L), and Sel (1.34 mg/L) allowed for the adaptation and proliferation of B. bovis without bovine serum, showed a constant increase in PPE, and reached a maximum value of 9.7% during seven cycles of in vitro culture. It was concluded that continuous proliferation of B. bovis in vitro could be achieved using A-DMEM/F12 medium supplemented with Ins-Trans-Sel, without bovine serum. After adaptation for proliferation in serum-free medium, the B. bovis strain of parasites could have future use in the study of this economically important protozoan species that affects cattle.
Altangerel, Khukhuu; Sivakumar, Thillaiampalam; Battsetseg, Badgar; Battur, Banzragch; Ueno, Akio; Igarashi, Ikuo; Yokoyama, Naoaki
We conducted a molecular epidemiological study on Babesia bovis in Mongolia. Three hundred blood samples collected from cattle grazed in seven different districts were initially screened using a previously established diagnostic polymerase chain reaction (PCR) assay for the detection of B. bovis-specific DNA. Positive samples were then used to amplify and sequence the hyper-variable regions of three B. bovis genes encoding the merozoite surface antigen (MSA)-1, MSA-2b, and MSA-2c. The diagnostic PCR assay detected B. bovis among cattle populations of all districts surveyed (4.4-26.0%). Sequences of each of the three genes were highly homologous among the Mongolian isolates, and found in a single phylogenetic cluster. In particular, a separate branch was formed only by the Mongolian isolates in the MSA-2b gene-based phylogenetic tree. Our findings indicate that effective preventative and control strategies are essential to control B. bovis infection in Mongolian cattle populations, and suggest that a careful approach must be adopted when using immunization techniques.
Asada, Masahito; Yahata, Kazuhide; Hakimi, Hassan; Yokoyama, Naoaki; Igarashi, Ikuo; Kaneko, Osamu
Genetic manipulation is an essential technique to analyze gene function; however, limited methods are available for Babesia bovis, a causative pathogen of the globally important cattle disease, bovine babesiosis. To date, two stable transfection systems have been developed for B. bovis, using selectable markers blasticidin-S deaminase (bsd) or human dihydrofolate reductase (hdhfr). In this work, we combine these two selectable markers in a sequential transfection system. Specifically, a parent transgenic B. bovis line which episomally expresses green fluorescent protein (GFP) and human dihydrofolate reductase (hDHFR), was transfected with a plasmid encoding a fusion protein consisting of red fluorescent protein (RFP) and blasticidin-S deaminase (BSD). Selection with WR99210 and blasticidin-S resulted in the emergence of parasites double positive for GFP and RFP. We then applied this method to complement gene function in a parasite line in which thioredoxin peroxidase-1 (Bbtpx-1) gene was knocked out using hDHFR as a selectable marker. A plasmid was constructed harboring both RFP-BSD and Bbtpx-1 expression cassettes, and transfected into a Bbtpx-1 knockout (KO) parasite. Transfectants were independently obtained by two transfection methods, episomal transfection and genome integration. Complementation of Bbtpx-1 resulted in full recovery of resistance to nitrosative stress, via the nitric oxide donor sodium nitroprusside, which was impaired in the Bbtpx-1 KO parasites. In conclusion, we developed a sequential transfection method in B. bovis and subsequently applied this technique in a gene complementation study. This method will enable broader genetic manipulation of Babesia toward enhancing our understanding of the biology of this parasite. PMID:25962142
Combrink, M P; Troskie, P C; de Klerk, D G; Pienaar, R; Latif, A A; Mans, B J
The South African Babesia bovis live blood vaccine, originating from a field isolate attenuated by 23 serial syringe passages in splenectomized calves, has lost the ability to infect the natural vector Rhipicephalus (Boophilus) microplus. In this study, infection with mixed parasites from the vaccine strain and a field isolate, resulted in transmission of both genotype populations. Comparing the field isolate and transmitted combination indicated no significant difference in their virulence, while challenge of vaccinated cattle with these isolates showed the ability of the vaccine to protect against both. Limiting dilution of the transmitted combination, followed by infection of splenectomized cattle (n=34) yielded no single infections for the vaccine strain genotype, seven clonal lines of the field isolate and one mixture of vaccine strain and field isolate. Only one of two field isolate clonal lines selected for vector transmission study was transmitted. Showing that B. bovis isolates can contain both tick transmissible and non-transmissible subpopulations. The findings of this study also indicate the probability of vaccine co-infection transmission occurring in the field, which may result in new genotype populations of B. bovis. However, the impact of this recombination with field isolates is considered negligible since a genotypically diverse population of B. bovis is already present in South Africa.
Nagano, Daisuke; Sivakumar, Thillaiampalam; De De Macedo, Alane Caine Costa; Inpankaew, Tawin; Alhassan, Andy; Igarashi, Ikuo; Yokoyama, Naoaki
In the present study, we screened blood DNA samples obtained from cattle bred in Brazil (n=164) and Ghana (n=80) for Babesia bovis using a diagnostic PCR assay and found prevalences of 14.6% and 46.3%, respectively. Subsequently, the genetic diversity of B. bovis in Thailand, Brazil and Ghana was analyzed, based on the DNA sequence of merozoite surface antigen-1 (MSA-1). In Thailand, MSA-1 sequences were relatively conserved and found in a single clade of the phylogram, while Brazilian MSA-1 sequences showed high genetic diversity and were dispersed across three different clades. In contrast, the sequences from Ghanaian samples were detected in two different clades, one of which contained only a single Ghanaian sequence. The identities among the MSA-1 sequences from Thailand, Brazil and Ghana were 99.0-100%, 57.5-99.4% and 60.3-100%, respectively, while the similarities among the deduced MSA-1 amino acid sequences within the respective countries were 98.4-100%, 59.4-99.7% and 58.7-100%, respectively. These observations suggested that the genetic diversity of B. bovis based on MSA-1 sequences was higher in Brazil and Ghana than in Thailand. The current data highlight the importance of conducting extensive studies on the genetic diversity of B. bovis before designing immune control strategies in each surveyed country.
Molad, T; Fleiderovitz, L; Leibovich, B; Wolkomirsky, R; Erster, O; Roth, A; Mazuz, M L; Markovics, A; Shkap, V
This study demonstrated the genetic diversity among MSA-2c, MSA-2a1 and MSA-2b proteins of Babesia bovis isolates obtained from bovine blood and Rhipicephalus annulatus tick samples. The least identities that were observed among the deduced amino acid sequences of MSA-2c, MSA-2a1 and MSA-2b were 55, 63, and 71%, respectively. During the study four B. bovis calves, aged about 1 month, were found to be infected with virulent field strains and developed babesiosis. Probably, the calves had received insufficient antibodies, or the antibodies raised against the vaccine strain did not cross-protect against virulent field isolates. The complete msa-2 locus from the Israeli B. bovis vaccine strain and two field isolates were characterized. Similarly to the Australian strains and isolates, the msa-2 loci of the examined Israeli strain and isolates had only two msa-2 genes - msa-2c and msa-2a/b - located between msa-2c and orfB. Several of the examined samples, contained different MSA-2 genotypes concurrently. No obvious geographical relationships among isolates from various regions of Israel were established. Moreover, in the phylogenetic analyses, the Israeli deduced MSA-2 amino acid sequences of the three examined genes were clustered together with sequences derived from other countries, proving that the msa-2 gene sequences of B. bovis shared the same genetic characters worldwide. The present study clearly showed that the MSA-2 proteins of B. bovis isolates from Israel were genetically distinct from the vaccine strains. Thus, further research will be needed in order to understand the genetic diversity mechanisms of B. bovis, and the immunological responses of the infected animals.
Jaramillo Ortiz, José Manuel; Del Médico Zajac, María Paula; Zanetti, Flavia Adriana; Molinari, María Paula; Gravisaco, María José; Calamante, Gabriela; Wilkowsky, Silvina Elizabeth
In this study, a recombinant modified vaccinia virus Ankara vector expressing a chimeric multi-antigen was obtained and evaluated as a candidate vaccine in homologous and heterologous prime-boost immunizations with a recombinant protein cocktail. The chimeric multi-antigen comprises immunodominant B and T cell regions of three Babesia bovis proteins. Humoral and cellular immune responses were evaluated in mice to compare the immunogenicity induced by different immunization schemes. The best vaccination scheme was achieved with a prime of protein cocktail and a boost with the recombinant virus. This scheme induced high level of specific IgG antibodies and secreted IFN and a high degree of activation of IFNγ(+) CD4(+) and CD8(+) specific T cells. This is the first report in which a novel vaccine candidate was constructed based on a rationally designed multi-antigen and evaluated in a prime-boost regime, optimizing the immune response necessary for protection against bovine babesiosis.
Gimenez, G; Belaunzarán, M L; Magalhães, K G; Poncini, C V; Lammel, E M; González Cappa, S M; Bozza, P T; Isola, E L D
Toll like receptors (TLRs) are involved in the modulation of diverse host genes expression through a complex network of signalling events that allow for an appropriate response to a microbial pathogen. In the present work we used TLR6KO mice in order to study the role of TLR6 in the immune discrimination of lipids from two Babesia bovis strains, attenuated R1A (LA) and virulent S2P (LV), and the consequent macrophage activation. We demonstrated that TLR6 is required for lipid body induction in murine peritoneal macrophages by both LA and LV. Interestingly, as regards IL-10 and COX-2/PGE2 pathway induction by LA and LV, we observed differences in the biological effects produced by these lipid extracts. Our results indicate a role of TLR6 in the down-modulation of these immunoregulators only in the case of LA, whereas this receptor was not implicated in pro-inflammatory TNFα, IL-6 and KC release induced by LA. Remarkably, LV did not exert the down-modulatory effect observed for LA, supporting the notion that LA and LV possess different lipid composition that could correlate with the polar pathogenic effect of both B. bovis strains.
Conservation of Babesia bovis small heat shock protein (Hsp20) among strains and definition of T helper cell epitopes recognized by cattle with diverse major histocompatibility complex class II haplotypes.
Norimine, Junzo; Mosqueda, Juan; Palmer, Guy H; Lewin, Harris A; Brown, Wendy C
Babesia bovis small heat shock protein (Hsp20) is recognized by CD4+ T lymphocytes from cattle that have recovered from infection and are immune to challenge. This candidate vaccine antigen is related to a protective antigen of Toxoplasma gondii, Hsp30/bag1, and both are members of the alpha-crystallin family of proteins that can serve as molecular chaperones. In the present study, immunofluorescence microscopy determined that Hsp20 is expressed intracellularly in all merozoites. Importantly, Hsp20 is also expressed by tick larval stages, including sporozoites, so that natural tick-transmitted infection could boost a vaccine-induced response. The predicted amino acid sequence of Hsp20 from merozoites is completely conserved among different B. bovis strains. To define the location of CD4+ T-cell epitopes for inclusion in a multiepitope peptide or minigene vaccine construct, truncated recombinant Hsp20 proteins and overlapping peptides were tested for their ability to stimulate T cells from immune cattle. Both amino-terminal (amino acids [aa] 1 to 105) and carboxy-terminal (aa 48 to 177) regions were immunogenic for the majority of cattle in the study, stimulating strong proliferation and IFN-gamma production. T-cell lines from all individuals with distinct DRB3 haplotypes responded to aa 11 to 62 of Hsp20, which contained one or more immunodominant epitopes for each animal. One epitope, DEQTGLPIKS (aa 17 to 26), was identified by T-cell clones. The presence of strain-conserved T helper cell epitopes in aa 11 to 62 of the ubiquitously expressed Hsp20 that are presented by major histocompatibility complex class II molecules represented broadly in the Holstein breed supports the inclusion of this region in vaccine constructs to be tested in cattle.
Apicomplexa tick borne hemoparasites including B. bovis, B. microti, and Theileria equi are responsible for bovine and human babesiosis and equine theileriosis respectively. These neglected parasites of vast medical, epidemiological, and economic impact have complex life cycles in their vertebrate a...
Babesia bovis is a tick-borne apicomplexan parasite that causes an acute disease in cattle. This study describes stable expression of an exogenous gfp-bsd gene in B. bovis transformed parasites. Cultured B. bovis infected erythrocytes of the biologically cloned Mo7 strain were transfected by electro...
Majid, A.A.; Bushera, H.O.; Saad, A.M.; Hussein, M.F.; Taylor, M.G.; Dargie, J.D.; Marshall, T.F.; Nelson, G.S.
Previous work has shown that cattle can acquire a strong resistance to Schistosoma bovis infection following repeated natural exposure. Partial resistance to a laboratory challenge with S. bovis has also been demonstrated in calves after immunization with an irradiated schistosomular or cercarial vaccine. The aim of the present study was to see whether this type of caccine could protect calves under the very different conditions of natural exposure to S. bovis in the field. Thirty 6- to 9-month-old calves were each immunized with 10,000 irradiated S. bovis schistosomula by intramuscular injection and 8 weeks later were released into an enzootic area along with 30 unvaccinated animals. The calves were followed up for 10 months, during which period protection was evidenced by a lower mortality rate, a slower rate of acquisition of infection, and lower fecal egg counts in the vaccinated calves. Necropsy of the survivors showed 60 to 70% reductions in worm and tissue egg counts of the vaccinated calves as compared to those not vaccinated.
Liu, Junlong; Guan, Guiquan; Li, Youquan; Liu, Aihong; Luo, Jianxun; Yin, Hong
Bovine babesiosis is a tick-transmitted disease caused by different species of Babesia. The white yak is a unique yak breed that lives only in Tianzhu in the Tibetan Autonomous County, Gansu Province, in northwestern China. Previous research using the ELISA method has confirmed that the white yak could become infected with B. bigemina. The objective of this study was the molecular detection and identification of Babesia species in white yaks. A total of 409 white yak blood samples were collected from 11 areas of the Tianzhu Tibetan Autonomous County in Northwest China from April to August, 2015. The V4 hypervariable region of Babesia 18S rRNA was amplified from extracted genomic DNA using nested PCR and sequenced. The nearly full-length sequence of 18S rRNA including the V4 region from the newly discovered Babesia was amplified and sequenced with Sanger method. PCR detection and sequencing indicated that 4/409 samples were positive for B. bigemina, 3/409 samples were positive for B. bovis, and 5/409 samples were positive for B. ovata. Additionally, a new Babesia species was found in 4/409 white yaks. A unique sequence of 1,627 bp was obtained from two of the four samples. The sequence was similar to Babesia species Akita (98.5%) found in Ixodes ovatus and B. venatorum (98%) and shared a 98% identity with B. divergens and a 98.1% identity with B. odocoilei. This study provides new data about Babesia infections in white yaks in northwestern China, and a new Babesia species similar to B. venatorum was identified in white yaks for the first time.
Liu, Junlong; Guan, Guiquan; Li, Youquan; Liu, Aihong; Luo, Jianxun; Yin, Hong
Bovine babesiosis is a tick-transmitted disease caused by different species of Babesia. The white yak is a unique yak breed that lives only in Tianzhu in the Tibetan Autonomous County, Gansu Province, in northwestern China. Previous research using the ELISA method has confirmed that the white yak could become infected with B. bigemina. The objective of this study was the molecular detection and identification of Babesia species in white yaks. A total of 409 white yak blood samples were collected from 11 areas of the Tianzhu Tibetan Autonomous County in Northwest China from April to August, 2015. The V4 hypervariable region of Babesia 18S rRNA was amplified from extracted genomic DNA using nested PCR and sequenced. The nearly full-length sequence of 18S rRNA including the V4 region from the newly discovered Babesia was amplified and sequenced with Sanger method. PCR detection and sequencing indicated that 4/409 samples were positive for B. bigemina, 3/409 samples were positive for B. bovis, and 5/409 samples were positive for B. ovata. Additionally, a new Babesia species was found in 4/409 white yaks. A unique sequence of 1,627 bp was obtained from two of the four samples. The sequence was similar to Babesia species Akita (98.5%) found in Ixodes ovatus and B. venatorum (98%) and shared a 98% identity with B. divergens and a 98.1% identity with B. odocoilei. This study provides new data about Babesia infections in white yaks in northwestern China, and a new Babesia species similar to B. venatorum was identified in white yaks for the first time. PMID:28352260
Sivakumar, Thillaiampalam; Tattiyapong, Muncharee; Fukushi, Shintaro; Hayashida, Kyoko; Kothalawala, Hemal; Silva, Seekkuge Susil Priyantha; Vimalakumar, Singarayar Caniciyas; Kanagaratnam, Ratnam; Meewewa, Asela Sanjeewa; Suthaharan, Kalpana; Puvirajan, Thamotharampillai; de Silva, Weligodage Kumarawansa; Igarashi, Ikuo; Yokoyama, Naoaki
Water buffaloes are thought to be the reservoir hosts for several hemoprotozoan parasites that infect cattle. In the present study, we surveyed Sri Lankan bred water buffaloes for infections with Babesia bovis, Babesia bigemina, Theileria annulata, and Theileria orientalis using parasite-specific PCR assays. When 320 blood-derived DNA samples from water buffaloes reared in three different districts (Polonnaruwa, Mannar, and Mullaitivu) of Sri Lanka were PCR screened, B. bovis, B. bigemina, and T. orientalis were detected. While T. orientalis was the predominant parasite (82.5%), low PCR-positive rates were observed for B. bovis (1.9%) and B. bigemina (1.6%). Amplicons of the gene sequences of the Rhoptry Associated Protein-1 (RAP-1) of B. bovis, the Apical Membrane Antigen-1 (AMA-1) of B. bigemina, and the Major Piroplasm Surface Protein (MPSP) of T. orientalis were compared with those characterized previously in Sri Lankan cattle. While the B. bigemina AMA-1 sequences from water buffaloes shared high identity values with those from cattle, B. bovis RAP-1 sequences from water buffaloes diverged genetically from those of cattle. For T. orientalis, none of the MPSP sequence types reported previously in Sri Lankan cattle (types 1, 3, 5, and 7) were detected in the water buffaloes, and the MPSP sequences analyzed in the present study belonged to types N1 or N2. In summary, in addition to reporting the first PCR-based survey of Babesia and Theileria parasites in water buffaloes in Sri Lanka, the present study found that the predominant variants of water buffalo-derived B. bovis RAP-1 and T. orientalis MPSP sequences were different from those previously described from cattle in this country.
Altangerel, Khukhuu; Alhassan, Andy; Iseki, Hiroshi; Sivakumar, Thillaiampalam; Boldbaatar, Damdinsuren; Yokoyama, Naoaki; Igarashi, Ikuo
A truncated fragment of the gene encoding the 200-kDa protein (P200) of Babesia bigemina was cloned into a plasmid vector, pGEX-4 T-1 and expressed in Escherichia coli as a glutathione-S-transferase fused protein. An indirect enzyme-linked immunosorbent assay (ELISA) using the rp200/CT detected specific antibodies in cattle experimentally infected with B. bigemina. Furthermore, the antigen did not cross-react with antibodies to Babesia bovis, a closely related Babesia parasite indicating that rp200/CT is a specific antigen for the diagnosis of B. bigemina infection. Additionally, ELISA using p200/CT and polymerase chain reaction were conducted on serum and corresponding DNA samples obtained from field cattle to evaluate the diagnostic utility of the p200/CT antigen. Results from the current study suggest that p200/CT ELISA is a sensitive and specific method for improved serodiagnosis of B. bigemina infection.
Bawm, Saw; Htun, Lat Lat; Maw, Ni Ni; Ngwe, Tin; Tosa, Yusuke; Kon, Tomoyuki; Kaneko, Chiho; Nakao, Ryo; Sakurai, Tatsuya; Kato, Hirotomo; Katakura, Ken
Cattle babesiosis is one of the most important tick-borne diseases worldwide. The present study reports a molecular survey of Babesia infections in cattle in Myanmar. Nested PCR assays based on the Babesia bigemina apical membrane antigen-1 gene (AMA-1) and B. bovis rhoptry associated protein-1 gene (RAP-1) revealed that the overall percentage of B. bigemina and B. bovis infection were 9.8% (70/713) and 17.1% (122/713), respectively. A mixed infection was detected in 4.6% (33/713) of animals. Animals <1 year (OR=13.66, CI=5.15-36.26) and 1-5 years of age (OR=3.91, CI=1.50-10.17) were identified as potential risk factors for B. bigemina infection. For B. bovis infection, age <1 year (OR=3.06, CI=1.63-5.75) and 1-5 years (OR=2.08, CI=1.21-3.57), Friesian-Zebu crossbreeds (OR=2.04, CI=1.26-3.30) and grazing (OR=1.59, CI=1.06-2.38) were identified as potential risk factors. This is the first report on a nationwide survey of bovine Babesia infections in Myanmar, providing useful information for the management and control of the disease.
Elsify, Ahmed; Sivakumar, Thillaiampalam; Nayel, Mohammed; Salama, Akram; Elkhtam, Ahmed; Rizk, Mohamed; Mosaab, Omar; Sultan, Khaled; Elsayed, Shimaa; Igarashi, Ikuo; Yokoyama, Naoaki
Cattle, buffaloes, and sheep are the main sources of meat and milk in Egypt, but their productivity is thought to be greatly reduced by hemoprotozoan parasitic diseases. In this study, we analyzed the infection rates of Babesia bovis, Babesia bigemina, Theileria annulata, and Theileria orientalis, using parasite-specific PCR assays in blood-DNA samples sourced from cattle (n=439), buffaloes (n=50), and sheep (n=105) reared in Menoufia, Behera, Giza, and Sohag provinces of Egypt. In cattle, the positive rates of B. bovis, B. bigemina, T. annulata, and T. orientalis were 3.18%, 7.97%, 9.56%, and 0.68%, respectively. On the other hand, B. bovis and T. orientalis were the only parasites detected in buffaloes and each of these parasites was only found in two individual DNA samples (both 2%), while one (0.95%) and two (1.90%) of the sheep samples were positive for B. bovis and B. bigemina, respectively. Sequence analysis showed that the B. bovis Rhoptry Associated Protein-1 and the B. bigemina Apical Membrane Antigen-1 genes were highly conserved among the samples, with 99.3-100% and 95.3-100% sequence identity values, respectively. In contrast, the Egyptian T. annulata merozoite surface antigen-1 gene sequences were relatively diverse (87.8-100% identity values), dispersing themselves across several clades in the phylogenetic tree containing sequences from other countries. Additionally, the T. orientalis Major Piroplasm Surface Protein (MPSP) gene sequences were classified as types 1 and 2. This is the first report of T. orientalis in Egypt, and of type 2 MPSP in buffaloes. Detection of MPSP type 2, which is considered a relatively virulent genotype, suggests that T. orientalis infection may have veterinary and economic significance in Egypt. In conclusion, the present study, which analyzed multiple species of Babesia and Theileria parasites in different livestock animals, may shed an additional light on the epidemiology of hemoprotozoan parasites in Egypt.
Jirapattharasate, Charoonluk; Adjou Moumouni, Paul Franck; Cao, Shinuo; Iguchi, Aiko; Liu, Mingming; Wang, Guanbo; Zhou, Mo; Vudriko, Patrick; Changbunjong, Tanasak; Sungpradit, Sivapong; Ratanakorn, Parntep; Moonarmart, Walasinee; Sedwisai, Poonyapat; Weluwanarak, Thekhawet; Wongsawang, Witsanu; Suzuki, Hiroshi; Xuan, Xuenan
Beef cattle production represents the largest cattle population in Thailand. Their productivity is constrained by tick-borne diseases such as babesiosis and theileriosis. In this study, we determined the prevalence of Babesia bigemina, Babesia bovis and Theileria orientalis using polymerase chain reaction (PCR). The genetic markers that were used for detection of the above parasites were sequenced to determine identities and similarity for Babesia spp. and genetic diversity of T. orientalis. Furthermore the risk factors for the occurrence of the above protozoan parasites in beef cattle from northern and northeastern parts of Thailand were assessed. A total of 329 blood samples were collected from beef cattle in 6 provinces. The study revealed that T. orientalis was the most prevalent (30.1%) parasite in beef cattle followed by B. bigemina (13.1%) and B. bovis (5.5%). Overall, 78.7% of the cattle screened were infected with at least one of the above parasites. Co-infection with Babesia spp. and T. orientalis was 30.1%. B. bigemina and T. orientalis were the most prevalent (15.1%) co-infection although triple infection with the three parasites was observed in 3.0% of the samples. Sequencing analysis revealed that B. bigemina RAP1 gene and B. bovis SBP2 gene were conserved among the parasites from different cattle samples. Phylogenetic analysis showed that the T. orientalis MPSP gene from parasites isolated from cattle in north and northeast Thailand was classified into types 5 and 7 as reported previously. Lack of tick control program was the universal risk factor of the occurrence of Babesia spp. and T. orientalis infection in beef cattle in northern and northeastern Thailand. We therefore recommend training of farmers on appropriate tick control strategies and further research on potential vectors for T. orientalis and elucidate the effect of co-infection with Babesia spp. on the pathogenicity of T. orientalis infection on beef in northern and northeastern Thailand.
Omar, Mosaab A; Salama, Akram; Elsify, Ahmed; Rizk, Mohamed Abdo; Al-Aboody, Mohammad Saleh; AbouLaila, Mahmoud; El-Sayed, Shimaa Abd El-Salam; Igarashi, Ikuo
Enoxacin is a broad-spectrum 6-fluoronaphthyridinone antibacterial agent (fluoroquinolones) structurally related to nalidixic acid used mainly in the treatment of urinary tract infections and gonorrhea. Also it has been shown recently that it may have cancer inhibiting effect. The primary antibabesial effect of Enoxacin is due to inhibition of DNA gyrase subunit A, and DNA topoisomerase. In the present study, enoxacin was tested as a potent inhibitor against the in vitro growth of bovine and equine Piroplasms. The in vitro growth of five Babesia species that were tested was significantly inhibited (P < 0.05) by micro molar concentrations of enoxacin (IC50 values = 33.5, 15.2, 7.5 and 23.2 μM for Babesia bovis, Babesia bigemina, Babesia caballi, and Theileria equi, respectively). Enoxacin IC50 values for Babesia and Theileria parasites were satisfactory as the drug is potent antibacterial drug with minimum side effects. Therefore, enoxacin might be used for treatment of Babesiosis and Theileriosis especially in case of mixed infections with bacterial diseases or incase of animal sensitivity against diminazin toxicity.
BEI Resources was developed by NIAID as a centralized biological resource center for research reagents to the scientific community (http://www.beiresources.org/). They have a considerable amount of reagents and isolates for parasitologists working with Entamoeba histolytica, Giardia, Toxoplasma, and...
The Merozoite Surface Antigens-2 of Babesia bovis conform a family of GPI-anchored glycoproteins located at the parasite cell surface, that contain neutralization-sensitive B-cell epitopes, thus constituting putative vaccine candidates for bovine babesiosis. It was previously shown that (i) the MSA-...
YAMASAKI, Masahiro; TSUBOI, Yoshihiro; TANIYAMA, Yusuke; UCHIDA, Naohiro; SATO, Reeko; NAKAMURA, Kensuke; OHTA, Hiroshi; TAKIGUCHI, Mitsuyoshi
The Babesia gibsoni heat shock protein 90 (BgHSP90) gene was cloned and sequenced. The length of the gene was 2,610 bp with two introns. This gene was amplified from cDNA corresponding to full length coding sequence (CDS) with an open reading frame of 2,148 bp. A phylogenetic analysis of the CDS of HSP90 gene showed that B. gibsoni was most closely related to B. bovis and Babesia sp. BQ1/Lintan and lies within a phylogenetic cluster of protozoa. Moreover, mRNA transcription profile for BgHSP90 exposed to high temperature were examined by quantitative real-time reverse transcription-polymerase chain reaction. BgHSP90 levels were elevated when the parasites were incubated at 43°C for 1 hr. PMID:27149891
Acosta, Fermín; Chernyaeva, Ekatherina; Mendoza, Libardo; Sambrano, Dilcia; Correa, Ricardo; Rotkevich, Mikhail; Tarté, Miroslava; Hernández, Humberto; Velazco, Bredio; de Escobar, Cecilia; de Waard, Jacobus H.
Panama remains free of zoonotic tuberculosis caused by Mycobacterium bovis. However, DNA fingerprinting of 7 M. bovis isolates from a 2013 bovine tuberculosis outbreak indicated minimal homology with strains previously circulating in Panama. M. bovis dispersion into Panama highlights the need for enhanced genotype testing to track zoonotic infections. PMID:25988479
Research conducted at the USDA/ARS/National Animal Disease Center in Ames, Iowa, reveals that ELISAs designed for detection of M. bovis-specific serum IgG in cattle may not be optimal for identification of seropositive bison, particularly those with low to moderate levels of antibody. In a study so...
Mycoplasma bovis in bison is a newly emerging and potentially devastating threat to the bison industry. This bacterium is increasingly being identified, both in the United States and Canada, as the cause of severe respiratory disease outbreaks with devastating consequences for the health of the ani...
el Bahrawy, A F; Nafei, S M; Morsy, T A; Farrag, A M
Babesiosis is a protozoal disease caused by members of the genus Babesia transmitted by the ixodid ticks. It is a parasite of various mammalian hosts as bovine (B. bovis), equine (B. equi), rodent (B. microti), canine (B. canis) and others. Human cases of babesiosis have been reported from several countries including Egypt. It is now well established that man may become infected (Fulminating or Subclinical) with several species of Babesia without prior splenectomy. In this paper, a total of 398 Rattus rattus and 90 R. norvegicus were trapped in Suez, Ismailia, Port Said, North Sinai and South Sinai Governorates. Blood films were taken from the atil after a simple cut of its end. The films were fixed in acetone free methyl alcohol and stained with Giemsa stain as usual. The results showed that all rats trapped in Ismailia had Babesia infection, then decreased in North Sinai, Port Said, South Sinai and was zero in Suez. As double infection. Trypanosoma lewisi was found in rats trapped in Port Said, North Sinai and South Sinai. But none had Theileria infection. The medical and veterinary importance of these results were discussed.
Munkhjargal, Tserendorj; AbouLaila, Mahmoud; Terkawi, Mohamad Alaa; Sivakumar, Thillaiampalam; Ichikawa, Madoka; Davaasuren, Batdorj; Nyamjargal, Tserendorj; Yokoyama, Naoaki; Igarashi, Ikuo
We evaluated the inhibitory effects of pepstatin A and mefloquine on the in vitro and in vivo growths of Babesia parasites. The in vitro growth of Babesia bovis, B. bigemina, B. caballi, and B. equi was significantly inhibited (P < 0.05) by micromolar concentrations of pepstatin A (50% inhibitory concentrations = 38.5, 36.5, 17.6, and 18.1 μM, respectively) and mefloquine (50% inhibitory concentrations = 59.7, 56.7, 20.7, and 4 μM, respectively). Furthermore, both reagents either alone at a concentration of 5 mg/kg or in combinations (2.5/2.5 and 5/5 mg/kg) for 10 days significantly inhibited the in vivo growth of B. microti in mice. Mefloquine treatment was highly effective and the combination treatments were less effective than other treatments. Therefore, mefloquine may antagonize the actions of pepstatin A against babesiosis and aspartic proteases may play an important role in the asexual growth cycle of Babesia parasites.
Tuvshintulga, Bumduuren; AbouLaila, Mahmoud; Davaasuren, Batdorj; Ishiyama, Aki; Sivakumar, Thillaiampalam; Yokoyama, Naoaki; Iwatsuki, Masato; Otoguro, Kazuhiko; Ōmura, Satoshi
The present study evaluated the growth-inhibitory effects of clofazimine, currently used for treating leprosy, against Babesia bovis, B. bigemina, B. caballi, and Theileria equi in in vitro culture and against Babesia microti in mice. The 50% inhibitory concentrations (IC50s) of clofazimine against the in vitro growth of B. bovis, B. bigemina, B. caballi, and T. equi were 4.5, 3, 4.3, and 0.29 μM, respectively. In mice infected with B. microti, treatment with 20 mg/kg of body weight of clofazimine administered orally resulted in a significantly lower peak parasitemia (5.3%) than that in the control group (45.9%), which was comparable to the subcutaneous administration of 25 mg/kg diminazene aceturate, the most widely used treatment for animal piroplasmosis. Although slight anemia was observed in both clofazimine- and diminazene aceturate-treated infected mice, the level and duration of anemia were lower and shorter, respectively, than those in untreated infected mice. Using blood transfusions and PCR, we also examined whether clofazimine completely killed B. microti. On day 40 postinfection, when blood analysis was performed, parasites were not found in blood smears; however, the DNA of B. microti was detected in the blood of clofazimine-treated animals and in several tissues of clofazimine- and diminazene aceturate-treated mice by PCR. The growth of parasites was observed in mice after blood transfusions from clofazimine-treated mice. In conclusion, clofazimine showed excellent inhibitory effects against Babesia and Theileria in vitro and in vivo, and further study on clofazimine is required for the future development of a novel chemotherapy with high efficacy and safety against animal piroplasmosis and, possibly, human babesiosis. PMID:26883713
Karlsson, Maria E; Andersson, Martin O
Babesiosis is an emerging tick-transmitted zoonosis in large parts of the world. In Sweden, the occurrence and diversity of Babesia species is largely unknown. In order to estimate the exposure to Babesia from infected ticks, we collected questing Ixodes ricinus from several sites across southern Sweden during two consecutive field seasons and investigated the occurrence of Babesia species. We report for the first time the occurrence of the zoonotic species Babesia venatorum in Swedish ticks, with a prevalence of 1%. We also detected B. microti (prevalence 3.2%) and B. divergens (prevalence 0.2%). The incidence of Babesia in questing ticks is substantially lower than that of several other tick-borne diseases in Sweden. Nevertheless, babesiosis should not be neglected as a possible diagnosis following tick bites in humans and animals in Sweden.
Ionita, Mariana; Mitrea, Ioan Liviu; Pfister, Kurt; Hamel, Dietmar; Buzatu, Catalin Marius; Silaghi, Cornelia
Canine babesiosis is a tick-borne disease caused by the protozoa Babesia spp. that affects dogs worldwide. In Romania, canine babesiosis has become quite frequent in the last few years, with a wide variety of clinical signs, ranging from mild, nonspecific illness to peracute collapse, and even death. Traditionally, a Babesia infection in dogs is diagnosed based on the morphologic appearance of the intraerythrocytic piroplasms observed in peripheral blood smears. To date, no data on genetic characterization of Babesia species in dogs has been documented for Romania. Therefore, a molecular survey on natural Babesia infections of dogs in Romania using polymerase chain reaction and genetic sequence analysis of a fragment of the ssRNA gene was performed. A total number of 16 blood samples were tested for the presence of Babesia DNA. Blood samples were collected from 11 dogs with symptoms of babesiosis and microscopically proven positive for Babesia and from a group of five asymptomatic dogs, not tested microscopically for Babesia, which were included in the study for comparative analysis. The piroplasm-specific PCR amplifying the partial 18S rRNA gene confirmed Babesia spp. infection in all 11 samples from dogs with clinical babesiosis, and in one of the clinically normal dogs. Sequence analysis revealed the presence of Babesia canis in all clinically affected dogs and Babesia vogeli in one clinically normal dog. This is the first molecular evidence of B. canis and B. vogeli in dogs from Romania. The results of the study provide basic information toward a better understanding of the epidemiology of canine babesiosis in Romania and will help to promote an effective control program.
da Silveira, Júlia A G; Rabelo, Elida M L; Ribeiro, Múcio F B
Intraerythrocytic protozoan species of the genera Theileria and Babesia are known to infect both wild and domestic animals, and both are transmitted by hard-ticks of the family Ixodidae. The prevalences of hemoprotozoa and ectoparasites in 15 free-living Mazama gouazoubira, two captive M. gouazoubira and four captive Blastocerus dichotomus from the State of Minas Gerais, Brazil, have been determined through the examination of blood smears and the use of nested polymerase chain reaction (nPCR). The cervid population was inspected for the presence of ticks and any specimens encountered were identified alive under the stereomicroscope. Blood samples were collected from all 21 animals, following which blood smears were prepared, subjected to quick Romanowsky staining and examined under the optical microscope. DNA was extracted with the aid of commercial kits from cervid blood samples and from tick salivary glands. The nPCR assay comprised two amplification reactions: the first was conducted using primers specific for a 1700 bp segment of the 18S rRNA gene of Babesia and Theileria species, whilst the second employed primers designed to amplify a common 420 bp Babesia 18S rRNA fragment identified by aligning sequences from Babesia spp. available at GenBank. The ticks Amblyomma cajennense, Rhipicephalus microplus and Dermacentor nitens were identified in various of the cervids examined. Of the animals investigated, 71.4% (15/21) were infected with hemoprotozoa, including Theileria cervi (47.6%), Theileria sp. (14.3%), Babesia bovis (4.8%) and Babesia bigemina (4.8%). However, only one of the infected wild cervids exhibited accentuated anaemia (PCV=17%). This is first report concerning the occurrence of Theileria spp. in Brazilian cervids.
Jackson, Andrew P.; Otto, Thomas D.; Darby, Alistair; Ramaprasad, Abhinay; Xia, Dong; Echaide, Ignacio Eduardo; Farber, Marisa; Gahlot, Sunayna; Gamble, John; Gupta, Dinesh; Gupta, Yask; Jackson, Louise; Malandrin, Laurence; Malas, Tareq B.; Moussa, Ehab; Nair, Mridul; Reid, Adam J.; Sanders, Mandy; Sharma, Jyotsna; Tracey, Alan; Quail, Mike A.; Weir, William; Wastling, Jonathan M.; Hall, Neil; Willadsen, Peter; Lingelbach, Klaus; Shiels, Brian; Tait, Andy; Berriman, Matt; Allred, David R.; Pain, Arnab
Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5′ ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct. PMID:24799432
Members of the CCp protein family have been previously described to be expressed on gametocytes of apicomplexan Plasmodium parasites. Knocking out Plasmodium CCp genes blocks the development of the parasite in the mosquito vector, making the CCp proteins potential targets for the development of a tr...
Blaschitz, Marion; Narodoslavsky-Gföller, Melanie; Kanzler, Michaela; Stanek, Gerold; Walochnik, Julia
Babesiosis is a tick-transmitted disease of veterinary and medical importance. The first Austrian case of human babesiosis was recently recorded. In the current study, ticks at all life cycle stages (instars), including 853 Ixodes ricinus and 11 Haemaphysalis concinna ticks, from sampling sites throughout Austria were examined for the presence of Babesia spp. by using 18S rRNA gene PCR and sequencing, and the overall mean infection rate was 51.04%. The infection rates for sampling sites were highly variable, ranging from 0% to almost 100%. Different instars and different sexes were infected almost equally. Babesia isolates occurring in Austrian ticks were identified as Babesia divergens, Babesia divergens-like, and Babesia sp. strain DD by sequencing a fragment of the heat shock protein 70 gene and internal transcribed spacer regions 1 and 2. To our knowledge, this is the first investigation of Babesia spp. in Austrian ticks. PMID:18539787
Background Human babesiosis is a rare tick-borne infectious disease. The clinical presentation ranges from an asymptomatic form to a life threatening infection with severe hemolysis. Human babesiosis due to Babesia microti is the most common and is endemic in North America. Case presentation We report a European patient with severe pancytopenia and reactive hemophagocytosis related to a Babesia microti infection. Babesia infection was acquired during a travel in the USA. Conclusion Babesiosis should be considered in patients who traveled in endemic areas, especially North America for the most common agent Babesia microti. PMID:23432953
Mørch, K; Holmaas, G; Frolander, P S; Kristoffersen, E K
Human babesiosis is a rare but potentially life-threatening parasitic disease transmitted by ixodid ticks, and has not previously been reported in Norway. We report a case of severe babesiosis that occurred in Norway in 2007. The patient had previously undergone a splenectomy. He was frequently exposed to tick bites in an area endemic for bovine babesiosis in the west of Norway. The patient presented with severe haemolysis and multiorgan failure. Giemsa-stained blood smears revealed 30% parasitaemia with Babesia spp. He was treated with quinine in combination with clindamycin, apheresis, and supportive treatment with ventilatory support and haemofiltration, and made a complete recovery. This is the first case reported in Norway; however Babesia divergens seroprevalence in cattle in Norway is high, as is the risk of Ixodes ricinus tick bite in the general population. Babesiosis should be considered in the differential diagnosis of unexplained febrile haemolytic disease.
The apicoplast is a highly specialized organelle that mediates required functions in the growth and replication of apicomplexan parasites. Despite structural conservation of the apicoplast among different parasite genera and species, there are also critical differences in the metabolic requirements ...
Arthropod borne pathogens have a complex life cycle that includes asexual reproduction of haploid stages in mammalian erythrocytes and development of diploid stages in the vector. Transition of Apicomplexan pathogens between the mammalian host and the arthropod vector is critical for ongoing transmi...
The apical complex of intracellular hemoparasites contains organelles like micronemes and rhoptries, specialized structures required for adherence and invasion of host cells. Several molecules discharged from rhoptries have been identified from Plasmodium spp., but only a single rhoptry associated p...
Figueroa, J V; Chieves, L P; Johnson, G S; Buening, G M
A SpeI-AvaI fragment (0.3 kbp) from pBbi16 (a pBR322 derivative containing a 6.3-kbp Babesia bigemina DNA insert) was subcloned into the pBluescript phagemid vector and was sequenced by the dideoxy-mediated chain termination method. Two sets of primers were designed for the polymerase chain reaction (PCR) assay. Primer set IA-IB was used to amplify a 278-bp DNA fragment, and primer set IAN-IBN was used to prepare a probe directed to a site within the PCR-amplified target DNA. Digoxigenin-dUTP was incorporated into the probe during the amplification reaction. PCR amplification of target DNA obtained from in vitro-cultured B. bigemina and nucleic acid hybridization of amplified product with the nonradioactive DNA probe showed that a 278-bp fragment could be detected when as little as 100 fg of parasite genomic DNA was used in the assay. A fragment of similar size was amplified from genomic DNAs from several B. bigemina isolates but not from DNAs from Babesia bovis, Anaplasma marginale, or six species of bacteria or bovine leukocytes. Similarly, the PCR product could be detected in DNA samples purified from 200 microliters of blood with a parasitemia of as low as 1 in 10(8) cells and which contained an estimated 30 B. bigemina-infected erythrocytes. By a direct PCR method, B. bigemina DNA was amplified from 20 microliters of packed erythrocytes with a calculated parasitemia of 1 in 10(9) cells. With the analytical sensitivity level of the PCR-DNA probe assay, six cattle with inapparent, 11-month chronic B. bigemina infection were found to be positive. No PCR product was observed in bovine blood samples collected from a splenectomized, A. marginale-infected bovine, a 4-year chronic B. bovis-infected animal, or 20 uninfected cattle from Missouri which were subjected to amplification. The PCR-DNA probe assay was shown to be sensitive in detecting latently infected cattle. The specificity and high analytical sensitivity of the test provide valuable tools for performing
Stewart, Linda D.; McNair, James; McCallan, Lyanne; Thompson, Suzan; Kulakov, Leonid A.
This study describes the development and optimization of an immunomagnetic separation (IMS) method to isolate Mycobacterium bovis cells from lymph node tissues. Gamma-irradiated whole M. bovis AF2122/97 cells and ethanol-extracted surface antigens of such cells were used to produce M. bovis-specific polyclonal and monoclonal antibodies in rabbits and mice. They were also used to generate M. bovis-specific peptide ligands by phage display biopanning. The various antibodies and peptide ligands obtained were used to coat MyOne tosyl-activated Dynabeads (Life Technologies), singly or in combination, and evaluated for IMS. Initially, M. bovis capture from Middlebrook 7H9 broth suspensions (concentration range, 10 to 105 CFU/ml) was evaluated by IMS combined with an M. bovis-specific touchdown PCR. IMS-PCR results and, subsequently, IMS-culture results indicated that the beads with greatest immunocapture capability for M. bovis in broth were those coated simultaneously with a monoclonal antibody and a biotinylated 12-mer peptide. These dually coated beads exhibited minimal capture (mean of 0.36% recovery) of 12 other Mycobacterium spp. occasionally encountered in veterinary tuberculosis (TB) diagnostic laboratories. When the optimized IMS method was applied to various M. bovis-spiked lymph node matrices, it demonstrated excellent detection sensitivities (50% limits of detection of 3.16 and 57.7 CFU/ml of lymph node tissue homogenate for IMS-PCR and IMS-culture, respectively). The optimized IMS method therefore has the potential to improve isolation of M. bovis from lymph nodes and hence the diagnosis of bovine tuberculosis. PMID:22322353
Carletti, Tamara; Barreto, Carmo; Mesplet, Maria; Mira, Anabela; Weir, William; Shiels, Brian; Oliva, Abel Gonzalez; Schnittger, Leonhard; Florin-Christensen, Monica
Babesia ovis, a tick-transmitted intraerythrocytic protozoan parasite, causes severe infections in small ruminants from Southern Europe, Middle East, and Northern Africa. With the aim of finding potential targets for the development of control methods against this parasite, sequence analysis of its genome led to the identification of four putative cysteine proteases of the C1A family. Orthology between B. ovis, B. bovis, T. annulata, and T. parva sequences showed that each B. ovis C1A peptidase sequence clustered within one of the four ortholog groups previously reported for these piroplasmids. The ortholog of bovipain-2 of B. bovis and falcipain-2 of Plasmodium falciparum, respectively, was designated "ovipain-2" and further characterized. In silico analysis showed that ovipain-2 has the typical topology of papain-like cysteine peptidases and a highly similar predicted three dimensional structure to bovipain-2 and falcipain-2, suggesting susceptibility to similar inhibitors. Immunoblotting using antibodies raised against a recombinant form of ovipain-2 (r-ovipain-2) demonstrated expression of ovipain-2 in in vitro cultured B. ovis merozoites. By immunofluorescence, these antibodies reacted with merozoites and stained the cytoplasm of infected erythrocytes. This suggests that ovipain-2 is secreted by the parasite and could be involved in intra- and extracellular digestion of hemoglobin and/or cleavage of erythrocyte proteins facilitating parasite egress. A significant reduction in the percentage of parasitized erythrocytes was obtained upon incubation of B. ovis in vitro cultures with anti-r-ovipain-2 antibodies, indicating an important functional role for ovipain-2 in the intra erythrocytic development cycle of this parasite. Finally, studies of the reactivity of sera from B. ovis-positive and negative sheep against r-ovipain-2 showed that this protease is expressed in vivo, and can be recognized by host antibodies. The results of this study suggest that ovipain-2
Maeda, Hiroki; Hatta, Takeshi; Alim, M Abdul; Tsubokawa, Daigo; Mikami, Fusako; Matsubayashi, Makoto; Miyoshi, Takeharu; Umemiya-Shirafuji, Rika; Kawazu, Shin-ichiro; Igarashi, Ikuo; Mochizuki, Masami; Tsuji, Naotoshi; Tanaka, Tetsuya
Ticks are potent vectors of many deadly human and animal pathogens. Tick-borne babesiosis is a well-recognized malaria-like disease that occurs worldwide and recently has attracted increased attention as an emerging zoonosis. Although the proliferation of Babesia organisms is essential in the vectors, their detailed lifecycle with time information for migration in ticks remains unknown. A novel study model for the elucidation of the migration speed of Babesia parasites in their vector tick, Haemaphysalis longicornis, has been developed using an artificial feeding system with quantitative PCR method. The detectable DNA of Babesia parasites gradually disappeared in the tick midgut at 1 day post engorgement (DPE), and in contrary increased in other organs. The results indicated that the Babesia parasite passed the H. longicornis midgut within 24 hours post engorgement, migrated to the hemolymph, and then proliferated in the organs except the midgut. This time point may be an important curfew for Babesia parasites to migrate in the tick lumen. We also visualized the Babesia parasites in the experimentally infected ticks and in their eggs using IFAT for detecting their cytoskeletal structure, which suggested the successful tick infection and transovarial transmission of the parasite. This model will shed light on the further understanding of tick-Babesia interactions. PMID:27841321
Yabsley, Michael J.; Shock, Barbara C.
Babesiosis is an emerging zoonotic disease on all inhabited continents and various wildlife species are the principal reservoir hosts for zoonotic Babesia species. The primary vectors of Babesia are Ixodid ticks, with the majority of zoonotic species being transmitted by species in the genus Ixodes. Species of Babesia vary in their infectivity, virulence and pathogenicity for people. Various factors (e.g., increased interactions between people and the environment, increased immunosuppression, changes in landscape and climate, and shifts in host and vector species abundance and community structures) have led to an increase in tick-borne diseases in people, including babesiosis. Furthermore, because babesiosis is now a reportable disease in several states in the United States, and it is the most common blood transfusion-associated parasite, recognized infections are expected to increase. Because of the zoonotic nature of these parasites, it is essential that we understand the natural history (especially reservoirs and vectors) so that appropriate control and prevention measures can be implemented. Considerable work has been conducted on the ecology of Babesia microti and Babesia divergens, the two most common causes of babesiosis in the United States and Europe, respectively. However, unfortunately, for many of the zoonotic Babesia species, the reservoir(s) and/or tick vector(s) are unknown. We review the current knowledge regarding the ecology of Babesia among their reservoir and tick hosts with an emphasis of the role on wildlife as reservoirs. We hope to encourage the molecular characterization of Babesia from potential reservoirs and vectors as well from people. These data are necessary so that informed decisions can be made regarding potential vectors and the potential role of wildlife in the ecology of a novel Babesia when it is detected in a human patient. PMID:24533312
Mostowy, Serge; Inwald, Jackie; Gordon, Steve; Martin, Carlos; Warren, Rob; Kremer, Kristin; Cousins, Debby; Behr, Marcel A.
Though careful consideration has been placed towards genetic characterization of tubercle bacillus isolates causing disease in humans, those causing disease predominantly among wild and domesticated mammals have received less attention. In contrast to Mycobacterium tuberculosis, whose host range is largely specific to humans, M. bovis and “M bovis-like” organisms infect a broad range of animal species beyond their most prominent host in cattle. To determine whether strains of variable genomic content are associated with distinct distributions of disease, the DNA contents of M. bovis or M. bovis-like isolates from a variety of hosts were investigated via Affymetrix GeneChip. Consistent with previous genomic analysis of the M. tuberculosis complex (MTC), large sequence polymorphisms of putative diagnostic and biological consequence were able to unambiguously distinguish interrogated isolates. The distribution of deleted regions indicates organisms genomically removed from M. bovis and also points to structured genomic variability within M. bovis. Certain genomic profiles spanned a variety of hosts but were clustered by geography, while others associated primarily with host type. In contrast to the prevailing assumption that M. bovis has broad host capacity, genomic profiles suggest that distinct MTC lineages differentially infect a variety of mammals. From this, a phylogenetic stratification of genotypes offers a predictive framework upon which to base future genetic and phenotypic studies of the MTC. PMID:16159772
Vega Y Murguia, C.A.
An in vitro model for the continuous replication of Babesia bigemina was developed and this model was used to study the parasite's biology. Initially, infected erythrocytes from a calf inoculated with a strain of B. bigemina was suspended with normal bovine erythrocytes and the parasite propagated in vitro. The cultured organism was inoculated into another calf and reproduced the disease with typical signs. Babesia bigemina was reisolated in pure culture in vitro. The animal recovered after receiving specific treatment. A procedure was developed to cryopreserve infected erythrocytes and merozoites to initiate in vitro cultures. Homogeneous parasite populations were obtained by cloning by limiting dilution. Parasitic growth was detected between 16-28 days after dilutions were made. Three primary clones were selected for recloning. Infected erythrocytes from the original isolate nd the clones were concentrated by Percoll density gradients. Density values for paired and single infected cells were determined. Enzymatic content of concentrated infected cells was analyzed by starch gel electrophoresis. Enzymes LDH, GPI, and GDH were detected, but polymorphism among clones was not observed. The enzyme 6-PDG was not associated with the parasite. Separation of labelled proteins was done by SDS-PAGE. The separation patterns were similar for all samples. a 43 Kd polypeptide was detected in the B. bigemina culture supernatant.
... M. bovis by eating or drinking contaminated, unpasteurized dairy products. The pasteurization process, which destroys disease-causing ... might put people at increased risk include ranching, dairy farming, working in a slaughterhouse or as a ...
Brooks, B W; Barnum, D A
The biochemical and morphological characteristics of 104 strains of Corynebacterium bovis isolated from bovine milk samples and the C. bovis reference strain were found to be uniform. Valuable criteria for identification were presence of catalase and oxidase, production of acid from glucose and fructose and a requirement for enriched basal media. Six strains of human and three strains of bovine origin were found to be inconsistent with the reference strain. PMID:6722650
Palmer, M V
Mycobacterium bovis is the cause of tuberculosis in animals and sometimes humans. Many developed nations have long-standing programmes to eradicate tuberculosis in livestock, principally cattle. As disease prevalence in cattle decreases these efforts are sometimes impeded by passage of M. bovis from wildlife to cattle. In epidemiological terms, disease can persist in some wildlife species, creating disease reservoirs, if the basic reproduction rate (R0) and critical community size (CCS) thresholds are achieved. Recognized wildlife reservoir hosts of M. bovis include the brushtail possum (Trichosurus vulpecula) in New Zealand, European badger (Meles meles) in Great Britain and Ireland, African buffalo (Syncerus caffer) in South Africa, wild boar (Sus scrofa) in the Iberian Peninsula and white-tailed deer (Odocoileus virginianus) in Michigan, USA. The epidemiological concepts of R0 and CCS are related to more tangible disease/pathogen characteristics such as prevalence, pathogen-induced pathology, host behaviour and ecology. An understanding of both epidemiological and disease/pathogen characteristics is necessary to identify wildlife reservoirs of M. bovis. In some cases, there is a single wildlife reservoir host involved in transmission of M. bovis to cattle. Complexity increases, however, in multihost systems where multiple potential reservoir hosts exist. Bovine tuberculosis eradication efforts require elimination of M. bovis transmission between wildlife reservoirs and cattle. For successful eradication identification of true wildlife reservoirs is critical, as disease control efforts are most effective when directed towards true reservoirs.
Del Carmen Terrón, María; González-Camacho, Fernando; González, Luis Miguel; Luque, Daniel; Montero, Estrella
The invasive form of the apicomplexan parasite Babesia divergens, the free merozoite, invades the erythrocytes of host vertebrates, leading to significant pathology. Although invasion is an active process critical for parasite survival, it is not yet entirely understood. Using techniques to isolate the viable free merozoite, as well as electron microscopy, we undertook a detailed morphological study and explored the sub-cellular structure of the invasive B. divergens free merozoite after it had left the host cell. We examined characteristic apicomplexan features such as the apicoplast, the inner and discontinuous double membrane complex, and the apical complex; some aspects of erythrocyte entry by B. divergens were also defined by electron microscopy. This study adds to our understanding of B. divergens free merozoites and their invasion of human erythrocytes.
Mycoplasma bovis (M. bovis) has recently emerged as a significant health threat in bison and is an increasing concern and source of economic loss for producers. Clinical manifestations of infection documented in bison include pneumonia, respiratory distress and polyarthritis. The current study des...
Dantrakool, Anchalee; Somboon, Pradya; Hashimoto, Tetsuo; Saito-Ito, Atsuko
A new type of rodent babesia, which resembled Babesia microti but was phylogenetically placed closest, with the highest level of statistical support, to Babesia canis, a canine babesia, was identified in Thai Bandicota indica in Thai provinces to which malaria is endemic. Close watch should be kept on human babesiosis in Thailand. PMID:14766871
Lempereur, Laetitia; Beck, Relja; Fonseca, Isabel; Marques, Cátia; Duarte, Ana; Santos, Marcos; Zúquete, Sara; Gomes, Jacinto; Walder, Gernot; Domingos, Ana; Antunes, Sandra; Baneth, Gad; Silaghi, Cornelia; Holman, Patricia; Zintl, Annetta
The genera Babesia and Theileria (phylum Apicomplexa, order Piroplasmida) are mainly transmitted by Ixodid ticks in which the sexual part of their life cycle followed by sporogony takes place. They include protozoan parasites that infect erythrocytes of a variety of vertebrate hosts, including domestic and wild animals, with some Babesia spp. also infecting humans. Babesia sporozoites transmitted in the tick's saliva during the bloodmeal directly infect erythrocytes, where they asexually multiply to produce pear-shaped merozoites in the process of merogony; whereas a pre-erythrocytic schizogonic life stage in leukocytes is found in Theileria and precedes merogony in the erythrocytes. The wide spectrum of Babesia and Theileria species and their dissimilar characteristics with relation to disease severity, transmission, epidemiology, and drug susceptibility stress the importance of accurate detection of babesiosis and theileriosis and their causative agents. These guidelines review the main methods currently used for the detection of Babesia and Theileria spp. for diagnostic purposes as well as epidemiological studies involving their vertebrate hosts and arthropod vectors. Serological methods were not included once they did not indicate current infection but rather exposure.
Silveira, Júlia A G; de Oliveira, Cairo H S; Silvestre, Bruna T; Albernaz, Tatiana T; Leite, Rômulo C; Barbosa, José D; Oliveira, Carlos M C; Ribeiro, Múcio F B
Approximately 50% of buffalo herds in Brazil are located in Pará state in northern Brazil. There are several properties where cattle and buffalo live and graze together, and thus, buffalo pathogens may threaten the health of cattle and vice versa. Therefore, knowledge of infectious agents of buffalo is essential for maintaining healthy livestock. Clinical disease caused by Theileria and Babesia parasites in the Asian water buffalo is not common, although these animals may act as reservoir hosts, and the detection of these hemoparasites in buffaloes is as important as it is in cattle. Studies of the infection of buffaloes by hemoparasites in Brazil are scarce. The objective of the present study was to investigate the occurrence of Piroplasmida parasites in Asian water buffaloes in the state of Pará in the Amazon region of Brazil using nested PCR assays and phylogenetic analysis. The 18S rRNA gene and ITS complete region were amplified from DNA extracted from blood samples collected from 308 apparently healthy buffaloes bred on six properties in the state of Pará, Brazil. The prevalence of positive buffalo samples was 4.2% (13/308) for Theileria spp., 3.6% (11/308) for Babesia bovis and 1% (3/308) for Babesia bigemina. Animals infected with Theileria were detected in 50% (3/6) of the assessed properties. Phylogenetic analyses indicated that the Theileria species detected in this study were closely related to Theileria buffeli, Theileria orientalis and Theileria sinensis. To our knowledge, this is the first report of Theileria in Asian water buffaloes in the Americas. The majority of Theileria-positive buffaloes (11/13) belong to a property that has a history of animals presenting lymphoproliferative disease of unknown etiology. Therefore, the present research suggests that this disorder can be associated with Theileria infection in this property. Our results provide new insights on the distribution and biological aspects of hemoparasites transmissible from
Mycoplasma bovis (M. bovis) is an emerging bacterial pathogen that has caused severe disease among ranched bison (Bison bison) herds in North America. Unlike cattle, M. bovis in bison seems to be a primary pathogen, affecting animals in feedlots as well as breeding-age cows on pasture. Mortality r...
An attenuated Mycobacterium bovis RD1 knockout (Delta RD1) vaccine administered to calves at 2 weeks of age provided similar efficacy as M. bovis bacille Calmette Guerin (BCG) against low dose, aerosol challenge with virulent M. bovis at 3.5m of age. Approximately 4.5 months after challenge, both De...
Chastant-Maillard, S; Boulouis, H-J; Reynaud, K; Thoumire, S; Gandoin, C; Bouillin, C; Cordonnier, N; Maillard, R
Transplacental transmission of Bartonella spp. has been reported for rodents, but not for cats and has never been investigated in cattle. The objective of this study was to assess vertical transmission of Bartonella in cattle. Fifty-six cow-calf pairs were tested before (cows) and after (calves) caesarean section for Bartonella bacteremia and/or serology, and the cotyledons were checked for gross lesions and presence of the bacteria. None of the 29 (52%) bacteremic cows gave birth to bacteremic calves, and all calves were seronegative at birth. Neither placentitis nor vasculitis were observed in all collected cotyledons. Bartonella bovis was not detected in placental cotyledons. Therefore, transplacental transmission of B. bovis and multiplication of the bacteria in the placenta do not seem likely. The lack of transplacental transmission may be associated with the particular structure of the placenta in ruminants or to a poor affinity/agressiveness of B. bovis for this tissue.
Battsetseg, B; Xuan, X; Ikadai, H; Bautista, J L; Byambaa, B; Boldbaatar, D; Battur, B; Battsetseg, G; Batsukh, Z; Igarashi, I; Nagasawa, H; Mikami, T; Fujisaki, K
Ticks play an important role in human and veterinary medicine particularly due to their ability to transmit protozoan pathogens. In this study we have demonstrated that polymerase chain reaction (PCR) and nested PCR methods enabled detection of Babesia caballi and Babesia equi in field isolates of Dermacentor nuttalli adult ticks from Mongolia. Primers specific for 218 bp fragment merozoite antigen 1 (EMA-1) gene of B. equi successfully amplified products from all samples of D. nuttalli adult ticks while primers for the 430 bp fragment product from BC48 gene of B. caballi amplified products from seven of the 54 samples. Using PCR and nested PCR methods we have found mixed infections with B. equi and B. caballi in the tick vector. The amplified DNA fragment from D. nuttalli ticks was inserted into the EcoRV site of pBluescript SK and sequenced. The sequence of the 430 bp fragment was completely identical to the nucleotide sequence of the USDA strain of B. caballi. These results suggest that D. nuttalli may play an important role as a vector of both B. caballi and B. equi and also may be important in maintaining endemicity of equine piroplasmosis in Mongolia.
Montero, Estrella; González, Luis Miguel; Chaparro, Alberto; Benzal, Jesús; Bertellotti, Marcelo; Masero, José A; Colominas-Ciuró, Roger; Vidal, Virginia; Barbosa, Andrés
This is the first reported case of Babesia sp. in Antarctic penguins, specifically a population of Chinstrap penguins (Pygoscelis antarctica) in the Vapour Col penguin rookery in Deception Island, South Shetlands, Antarctica. We collected peripheral blood from 50 adult and 30 chick Chinstrap penguins. Examination of the samples by microscopy showed intraerythrocytic forms morphologically similar to other avian Babesia species in 12 Chinstrap penguin adults and seven chicks. The estimated parasitaemias ranged from 0.25×10(-2)% to 0.75×10(-2)%. Despite the low number of parasites found in blood smears, semi-nested PCR assays yielded a 274 bp fragment in 12 of the 19 positive blood samples found by microscopy. Sequencing revealed that the fragment was 97% similar to Babesia sp. 18S rRNA from Australian Little Penguins (Eudyptula minor) confirming presence of the parasite. Parasite prevalence estimated by microscopy in adults and chicks was higher (24% vs. 23.3%, respectively) than found by semi-nested PCR (16% vs. 13.3% respectively). Although sampled penguins were apparently healthy, the effect of Babesia infection in these penguins is unknown. The identification of Babesia sp. in Antarctic penguins is an important finding. Ixodes uriae, as the only tick species present in the Antarctic Peninsula, is the key to understanding the natural history of this parasite. Future work should address the transmission dynamics and pathogenicity of Babesia sp. in Chinstrap penguin as well as in other penguin species, such as Gentoo penguin (Pygoscelis papua) and Adélie penguin (Pygoscelis adeliae), present within the tick distribution range in the Antarctic Peninsula.
Reichard, Mason V; Gray, Kristene M; Van den Bussche, Ronald A; d'Offay, Jean M; White, Gary L; Simecka, Christine M; Wolf, Roman F
Babesia spp. are tick-transmitted apicomplexan hemoparasites that infect mammalian red blood cells. Our purpose was to determine the prevalence of Babesia infection in a colony of captive baboons and to evaluate potential experimental routes of the transmission of the hemoparasite. DNA was extracted from the blood of baboons and tested for infection with Babesia by PCR and primers that amplify the 18s rRNA gene of the parasite. The overall prevalence of infection of Babesia in the baboon population was 8.8% (73 of 830). Phylogenetic analysis of the sequenced DNA from 2 baboons revealed that the Babesia isolate found in captive baboons was a novel species most closely related (97% to 99%) to B. leo. Blood from a Babesia-infected donor baboon was inoculated intravenously, intramuscularly, or subcutaneously into 3 naive baboons. The intravenously inoculated baboon was PCR-positive at 7 d after inoculation; the 2 baboons inoculated by other routes became PCR-positive at 10 d after inoculation. All 3 baboons remained PCR-positive for Babesia through day 31. Baboons experimentally inoculated with the new Babesia isolate did not exhibit clinical signs of babesiosis during the experiments. We demonstrated that captive baboons are infected with a novel Babesia isolate. In addition we showed that Babesia can be transmitted in the absence of the organism's definitive host (ticks) by transfer of infected blood through intravenous, intramuscular, and subcutaneous routes to naive baboons.
Aerosol and intratracheal inoculation routes are commonly used for experimental biology purposes to infect cattle with virulent Mycobacterium bovis, each resulting primarily in a respiratory tract infection including lungs and lung-associated lymph nodes. Disease severity is dose and time dependent...
Arsuaga, Marta; Gonzalez, Luis M; Lobo, Cheryl A; de la Calle, Fernando; Bautista, Jose M; Azcárate, Isabel G; Puente, Sabino; Montero, Estrella
Babesiosis is an emerging zoonosis now found in several areas of the world. Using PCR and indirect immunofluorescence assay, we have diagnosed the first case of human babesiosis caused by Babesia microti in Spain. Diagnosis was delayed because of the nonspecific clinical symptoms that occurred in an immunocompetent patient.
Clark, Kerry; Savick, Kyla; Butler, Joseph
Human babesiosis in the United States is caused most commonly by the intraerythrocytic protozoan parasite, Babesia microti . Although a few reports have described evidence of Babesia species in animals in Florida, to date Babesia microti specifically has not been reported from Florida or most other southern states. To determine if the organism is present in vertebrates in the region, small mammals were trapped and sampled at 2 sites in northeastern Florida, and DNA extracts from blood samples were screened for B. microti DNA via PCR assays targeting portions of the nuclear small subunit rRNA (18S rDNA) and beta-tubulin genes. Amplified fragments from representative samples of PCR-positive hosts were sequenced and compared phylogenetically to reference strains of Babesia species. The B. microti strains found in cotton rats ( Sigmodon hispidus ) most closely resembles B. microti sensu stricto strains that are pathogenic to humans, and strains found in raccoons ( Procyon lotor ) most closely resembles previously described raccoon-related strains of B. microti sensu lato. The results of this study suggest that B. microti is prevalent among cotton rats and raccoons at some sites in northeast Florida and may pose a risk to humans in the region.
Arsuaga, Marta; Gonzalez, Luis M.; Lobo, Cheryl A.; de la Calle, Fernando; Bautista, Jose M.; Azcárate, Isabel G.; Puente, Sabino
Abstract Babesiosis is an emerging zoonosis now found in several areas of the world. Using PCR and indirect immunofluorescence assay, we have diagnosed the first case of human babesiosis caused by Babesia microti in Spain. Diagnosis was delayed because of the nonspecific clinical symptoms that occurred in an immunocompetent patient. PMID:27560451
Early interactions of innate immune cell populations such as DC, monocytes/macrophages and NK cells, can affect the ability of the acquired immune response to control infection of intracellular microorganisms. In this study, we investigated the activation of bovine NK cells by CD13+ splenic DC or CD...
Antunes, Sandra; Merino, Octávio; Lérias, Joana; Domingues, Nuno; Mosqueda, Juan; de la Fuente, José; Domingos, Ana
Ticks are obligate haematophagous ectoparasites considered the principal vectors of disease among animals. Rhipicephalus microplus and R. annulatus ticks are the most important vectors for Babesia bigemina and B. bovis, two of the most important intraerythrocytic protozoan parasites species in cattle, responsible for babesiosis which together with anaplasmosis account for substantial economic losses in the livestock industry worldwide. Anti-tick vaccines are a proved alternative to traditional tick and tick borne diseases control methods but are still limited primarily due to the lack of effective antigens. Subsequently to the identification of antigens the validation is a laborious work often expensive. Tick artificial feeding, is a low cost alternative to test antigens allowing achieving critical data. Herein, R. microplus females were successfully artificially fed using capillary tubes. Calreticulin (CRT) protein, which in a previous study has been identified as being involved in B. bigemina infection in R. annulatus ticks, was expressed as recombinant protein (rCRT) in an E. coli expression system and antibodies raised against rCRT. Anti-rCRT serum was supplemented to a blood meal, offered to partially engorged R. microplus females and their effect in feeding process as well as infection by B. bigemina was analyzed. No significant reductions in tick and egg weight were observed when ticks fed with anti-rCRT serum. Furthermore, B. bigemina infection levels did not show a statistically significant decrease when ticks fed with anti-rCRT antibodies. Results suggest that CRT is not a suitable candidate for cattle vaccination trials.
Duarte, Sabrina Castilho; Linhares, Guido Fontgalland Coelho; Romanowsky, Tatiana Nunes; da Silveira Neto, Osvaldo José; Borges, Ligia Miranda Ferreira
Canine babesiosis is an infectious disease caused by either Babesia gibsoni or Babesia canis protozoans. The latter is also classified under three different phylogenetic groups, referred to as subspecies B. canis canis, B. canis vogeli and B. canis rossi. The objective of the present study was to validate and standardize a PCR assay to discriminate the organisms at the subspecies level. First, the reference sequences of the 18S rRNA, 5.8S rRNA and 28S rRNA genes, including the internal transcribed spacer 1 (ITS1) and 2 (ITS2) of the most common species and subspecies of the genus Babesia were retrieved from the GenBank database. Subspecies-specific primers (BAB3, BAB4 and BAB5) and one genus-specific primer were designed from the alignment of the sequences. The PCR assays were evaluated in three different combinations of primer pairs in order to assure complete specificity for each reaction. The results of the tests had demonstrated effectiveness of the novel primer pairs BAB1/BAB3, BAB1/BAB4 and BAB1/BAB5 for the amplification of the subspecies-specific target fragments of 746 bp (B. c. canis), 546 bp (B. c. vogeli) and 342 bp (B. c. rossi) by PCR. The original enzymatic amplification assays with novel primers reported in this paper were confirmed to be a reliable tool for the specific discrimination among B. canis subspecies by single-step PCR assays.
Niu, Qingli; Liu, Zhijie; Yang, Jifei; Guan, Guiquan; Pan, Yuping; Luo, Jianxun; Yin, Hong
Apical membrane antigen-1 (AMA-1) has been described as a potential vaccine candidate in apicomplexan parasites. Here we characterize the ama-1 gene. The full-length ama-1 gene of Babesia sp. BQ1 (Lintan) (BLTAMA-1) is 1785 bp, which contains an open reading frame (ORF) encoding a 65-kDa protein of 594 amino acid residues; by definition, the 5' UTR precedes the first methionine of the ORF. Phylogenetic analysis based on AMA-1 amino acid sequences clearly separated Piroplasmida from other Apicomplexa parasites. The Babesia sp. BQ1 (Lintan) AMA-1 sequence is most closely associated with that of B. ovata and B. bigemina, with high bootstrap value. A recombinant protein encoding a conserved region and containing ectodomains I and II of BLTAMA-1 was constructed. BLTrAMA-1-DI/DII proteins were tested for reactivity with sera from sheep infected by Babesia sp. BQ1 (Lintan). In Western-blot analysis, native Babesia sp. BQ1 (Lintan) AMA-1 proteins were recognized by antibodies raised in rabbits against BLTrAMA-1 in vitro. The results of this study are discussed in terms of gene characterization, taxonomy and antigenicity.
Lysnyansky, Inna; Ayling, Roger D.
Mycoplasma bovis is a cell-wall-less bacterium and belongs to the class Mollicutes. It is the most important etiological agent of bovine mycoplasmoses in North America and Europe, causing respiratory disease, mastitis, otitis media, arthritis, and reproductive disease. Clinical disease associated with M. bovis is often chronic, debilitating, and poorly responsive to antimicrobial therapy, resulting in significant economic loss, the full extent of which is difficult to estimate. Until M. bovis vaccines are universally available, sanitary control measures and antimicrobial treatment are the only approaches that can be used in attempts to control M. bovis infections. However, in vitro studies show that many of the current M. bovis isolates circulating in Europe have high minimum inhibitory concentrations (MIC) for many of the commercially available antimicrobials. In this review we summarize the current MIC trends indicating the development of antimicrobial resistance in M. bovis as well as the known molecular mechanisms by which resistance is acquired. PMID:27199926
Yabsley, Michael J; Murphy, Staci M; Cunningham, Mark W
Piroplasms, morphologically indistinguishable from Cytauxzoon felis, previously were detected in 36% of cougars in Florida. We utilized a nested 18S rRNA assay, which amplifies DNA from all piroplasms, to screen blood samples collected from 41 cougars from Florida (39 native Florida panthers [Puma concolor coryi] and two translocated Texas cougars [P. c. stanleyana]) from 1989-2005. Thirty-nine of the 41 cougars (95%) were positive for piroplasms; however, sequence analysis and restriction enzyme digestion revealed that only five were positive for C. felis. Samples from 32 cougars were positive for a Babesia sp. Two cougars were co-infected with both C. felis and the Babesia sp. Phylogenetic analysis of 18S rRNA gene sequence indicated that the Florida panther Babesia sp. was most closely related to a Babesia sp. reported from Ixodes ovatus from Japan, Babesia divergens, and Babesia odocoilei. This study indicates that Florida panthers harbor two distinct piroplasms, C. felis and a Babesia sp., and that some individuals are infected with both organisms. The infectivity and pathogenicity of this Babesia sp. for domestic cats is unknown. This represents the first report of a feline Babesia sp. in North America.
Apicomplexan parasites commonly contain a unique, non-photosynthetic plastid-like organelle termed the apicoplast. Previous analyses of other plastid-containing organisms suggest that apicoplasts were derived from a red algal ancestor. In this report, we present an extensive phylogenetic study of ap...
The spleen is the critical organ in defense against hemoparasitic diseases like babesiosis, responding to the pathogen in blood as it passes through the organ. As a lymphoid organ, the spleen is composed of cells whose role it is to orchestrate the engagement of infected erythrocytes and free parasi...
Wong, Samson S. Y.; Poon, Rosana W. S.; Hui, Janet J. Y.
Intraerythrocytic Babesia-like trophozoites were seen in postmortem kidney sections of a free-roaming cat in Hong Kong. DNA sequences of the 18S rRNA and mitochondrial cytochrome b genes had only 96.7% and 90.4% nucleotide identity with known Babesia sequences. We propose that this new species be named Babesia hongkongensis. PMID:22649017
Brennan, Meghan B; Herwaldt, Barbara L; Kazmierczak, James J; Weiss, John W; Klein, Christina L; Leith, Catherine P; He, Rong; Oberley, Matthew J; Tonnetti, Laura; Wilkins, Patricia P; Gauthier, Gregory M
Babesia microti, an intraerythrocytic parasite, is tickborne in nature. In contrast to transmission by blood transfusion, which has been well documented, transmission associated with solid organ transplantation has not been reported. We describe parasitologically confirmed cases of babesiosis diagnosed ≈8 weeks posttransplantation in 2 recipients of renal allografts from an organ donor who was multiply transfused on the day he died from traumatic injuries. The organ donor and recipients had no identified risk factors for tickborne infection. Antibodies against B. microti parasites were not detected by serologic testing of archived pretransplant specimens. However, 1 of the organ donor's blood donors was seropositive when tested postdonation and had risk factors for tick exposure. The organ donor probably served as a conduit of Babesia parasites from the seropositive blood donor to both kidney recipients. Babesiosis should be included in the differential diagnosis of unexplained fever and hemolytic anemia after blood transfusion or organ transplantation.
Herwaldt, Barbara L.; Kazmierczak, James J.; Weiss, John W.; Klein, Christina L.; Leith, Catherine P.; He, Rong; Oberley, Matthew J.; Tonnetti, Laura; Wilkins, Patricia P.; Gauthier, Gregory M.
Babesia microti, an intraerythrocytic parasite, is tickborne in nature. In contrast to transmission by blood transfusion, which has been well documented, transmission associated with solid organ transplantation has not been reported. We describe parasitologically confirmed cases of babesiosis diagnosed ≈8 weeks posttransplantation in 2 recipients of renal allografts from an organ donor who was multiply transfused on the day he died from traumatic injuries. The organ donor and recipients had no identified risk factors for tickborne infection. Antibodies against B. microti parasites were not detected by serologic testing of archived pretransplant specimens. However, 1 of the organ donor’s blood donors was seropositive when tested postdonation and had risk factors for tick exposure. The organ donor probably served as a conduit of Babesia parasites from the seropositive blood donor to both kidney recipients. Babesiosis should be included in the differential diagnosis of unexplained fever and hemolytic anemia after blood transfusion or organ transplantation. PMID:27767010
Camacho, A T; Guitian, F J; Pallas, E; Gestal, J J; Olmeda, A S; Habela, M A; Telford, S R; Spielman, A
The control of equine piroplasmosis is becoming increasingly important to maintain the international market open to the horse industry. The purpose of this study was to demonstrate the occurrence of equine piroplasmosis (Theileria equi and Babesia caballi) in Galicia, north-west Spain, and to compare haematological and serum biochemistry parameters between non-parasitaemic horses and horses parasitaemic with T. equi and B. caballi. Sixty serum samples (control group) were taken from healthy horses pastured on two farms, and examined for evidence of equine T. equi and B. caballi infection by indirect fluorescent antibody test (IFAT). Of the 60 samples, 24 (40%) and 17 (28.3%) samples were positive for T. equi and B. caballi, respectively. Twelve (20%) samples were positive for both parasites. Haematology and serum biochemistry were compared between controls and a series of 36 horses clinically affected by T. equi (25) or B. caballi (11). Compared with the healthy group, there was a 43% and 37% decrease in the haematocrit for T. equi and B. caballi infection, respectively. Parasitaemic horses presented an intense anaemia and serum biochemistry signs of liver damage. The anaemia was more severe in T. equi-infected than in B. caballi-infected horses. Our results suggest that equine piroplasmosis is widespread in the region and is a cause for concern.
Tsuji, Naotoshi; Battsetseg, Badgar; Boldbaatar, Damdinsuren; Miyoshi, Takeharu; Xuan, Xuenan; Oliver, James H.; Fujisaki, Kozo
Antimicrobial peptides are major components of host innate immunity, a well-conserved, evolutionarily ancient defensive mechanism. Infectious disease-bearing vector ticks are thought to possess specific defense molecules against the transmitted pathogens that have been acquired during their evolution. We found in the tick Haemaphysalis longicornis a novel parasiticidal peptide named longicin that may have evolved from a common ancestral peptide resembling spider and scorpion toxins. H. longicornis is the primary vector for Babesia sp. parasites in Japan. Longicin also displayed bactericidal and fungicidal properties that resemble those of defensin homologues from invertebrates and vertebrates. Longicin showed a remarkable ability to inhibit the proliferation of merozoites, an erythrocyte blood stage of equine Babesia equi, by killing the parasites. Longicin was localized at the surface of the Babesia sp. parasites, as demonstrated by confocal microscopic analysis. In an in vivo experiment, longicin induced significant reduction of parasitemia in animals infected with the zoonotic and murine B. microti. Moreover, RNA interference data demonstrated that endogenous longicin is able to directly kill the canine B. gibsoni, thus indicating that it may play a role in regulating the vectorial capacity in the vector tick H. longicornis. Theoretically, longicin may serve as a model for the development of chemotherapeutic compounds against tick-borne disease organisms. PMID:17485458
Tsuji, Naotoshi; Battsetseg, Badgar; Boldbaatar, Damdinsuren; Miyoshi, Takeharu; Xuan, Xuenan; Oliver, James H; Fujisaki, Kozo
Antimicrobial peptides are major components of host innate immunity, a well-conserved, evolutionarily ancient defensive mechanism. Infectious disease-bearing vector ticks are thought to possess specific defense molecules against the transmitted pathogens that have been acquired during their evolution. We found in the tick Haemaphysalis longicornis a novel parasiticidal peptide named longicin that may have evolved from a common ancestral peptide resembling spider and scorpion toxins. H. longicornis is the primary vector for Babesia sp. parasites in Japan. Longicin also displayed bactericidal and fungicidal properties that resemble those of defensin homologues from invertebrates and vertebrates. Longicin showed a remarkable ability to inhibit the proliferation of merozoites, an erythrocyte blood stage of equine Babesia equi, by killing the parasites. Longicin was localized at the surface of the Babesia sp. parasites, as demonstrated by confocal microscopic analysis. In an in vivo experiment, longicin induced significant reduction of parasitemia in animals infected with the zoonotic and murine B. microti. Moreover, RNA interference data demonstrated that endogenous longicin is able to directly kill the canine B. gibsoni, thus indicating that it may play a role in regulating the vectorial capacity in the vector tick H. longicornis. Theoretically, longicin may serve as a model for the development of chemotherapeutic compounds against tick-borne disease organisms.
Peirce, M.A.; Parsons, N.J.
A new species of haematozoa, Babesia ugwidiensis sp. nov. from a cormorant is described. This is the first species of piroplasm to be recorded from the Phalacrocoracidae and the relationship of this parasite to other Babesia spp. from marine hosts is discussed. PMID:23193522
Puraite, Irma; Rosef, Olav; Radzijevskaja, Jana; Lipatova, Indre; Paulauskas, Algimantas
Babesiosis is an emerging zoonotic disease and various wildlife species are reservoir hosts for zoonotic species of Babesia Starcovici, 1893. The objective of the present study was to investigate the presence and prevalence of Babesia spp. in moose Alces alces (Linnaeus) in two regions of Norway. A total of 99 spleen samples were collected from animals of various ages from an area with the occurrence of the tick Ixodes ricinus (Linnaeus, 1758), and from an area where the ticks are known to be absent. Infection was detected by the amplification of different regions of the 18S rRNA gene by using two different PCR primer sets specific of Babesia. Babesia spp. were found in the spleen samples of four moose. All Babesia-infected animals were from an area where ticks occur, with an infection rate of 6% (4 of 70). Babesia-positive samples were obtained from a five-month old moose calf and three adults. Two Babesia species, Babesia capreoli (Enigk et Friedhoff, 1962) and a B. odocoilei-like, were identified. Co-infection with Anaplasma phagocytophilum was obtained in two animals. This is the first report of the occurrence of B. capreoli and B. odocoilei-like species in moose.
Jadeja, L.; Kantarjian, H.; Bolivar, R.
We describe the first patient with simultaneous S bovis septicemia and meningitis associated with chronic radiation enterocolitis. This case underlines the value of a thorough gastrointestinal evaluation of all patients with S bovis infection, and the need for a neurologic investigation even with minor neurologic manifestations.
Mycoplasma bovis (M. bovis) is an emerging, primary pathogen of ranched bison (Bison bison) in North America. It causes severe disease among animals in feedlots as well as breeding-age cows and bulls on pasture. Mortality in adult bison is as high as 25 percent, resulting in significant economic l...
In the early 20th century there were large numbers of tuberculous cattle in many countries. An association was made between the number of M. bovis infected humans and the prevalence of tuberculosis in cattle. Mandatory pasteurization of milk caused the prevalence of human tuberculosis due to M. bovi...
Objective. Mycobacterium bovis is the causative agent of bovine tuberculosis, a relevant zoonosis that can spread to humans through inhalation or by ingestion. M. bovis multiplies slowly, so infected animals may be sent to slaughter during the early stages of the disease before diagnosis and when ...
Dan, J M; Crespo, M; Silveira, F P; Kaplan, R; Aslam, S
We present a report of extrapulmonary Mycobacterium bovis infection in a lung transplant recipient. M. bovis is acquired predominantly by zoonotic transmission, particularly from consumption of unpasteurized foods. We discuss epidemiologic exposure, especially as relates to the Mexico-US border, clinical characteristics, resistance profile, and treatment.
Bosman, Anna-Mari; Oosthuizen, Marinda C; Peirce, Michael A; Venter, Estelle H; Penzhorn, Barend L
In a previous paper, we reported on a large number of cheetah blood specimens that gave positive signals only for Babesia and/or Theileria genus-specific probes on the reverse line blot (RLB) assay, indicating the presence of a novel species or variant of an existing species. Some of these specimens were investigated further by microscopic, serological, sequencing, and phylogenetic analyses. The near-full-length 18S rRNA genes of 13 samples, as well as the second internal transcribed spacer (ITS2) region, were amplified, cloned, and sequenced. A species-specific RLB probe, designed to target the hypervariable V4 region of the 18S rRNA gene for detection of the novel Babesia sp., was used to screen an additional 137 cheetah blood specimens for the presence of the species. The prevalence of infection was 28.5%. Here we describe the morphology and phylogenetic relationships of the novel species, which we have named Babesia lengau sp. nov.
Mol, J P S; Carvalho, T F; Fonseca, A A; Sales, E B; Issa, M A; Rezende, L C; Hodon, M A; Tinoco, H P; Malta, M C C; Pessanha, A T; Pierezan, F; Mota, P M P C; Paixão, T A; Santos, R L
Tuberculosis, associated with Mycobacterium bovis, was diagnosed post mortem in an adult female capybara (Hydrochoerus hydrochaeris), kept at the Pampulha Ecological Park, Belo Horizonte, Brazil, in a large metropolitan area. On post-mortem examination, there were numerous firm white nodules scattered throughout all lobes of both lungs. Tissue samples were collected for histological and microbiological examination. Microscopically, the pulmonary nodules were multifocal to coalescing granulomas and intralesional acid-fast bacilli were evident in Ziehl-Neelsen-stained sections of the lung and spleen. Colonies with morphological features of Mycobacterium spp. were isolated from lung samples and conventional polymerase chain reaction (PCR) with genomic DNA from the isolates was positive for M. bovis; sequencing indicated 100% identity with the region of difference 4 (RD4) of M. bovis. In addition, M. bovis DNA was detected in the lung by quantitative PCR. The finding of M. bovis in a capybara indicates a potential public health risk in a zoological collection.
Mycobacterium bovis (M. bovis) causes tuberculosis in white-tailed deer (WTD). Natural infection of WTD with M. bovis is most closely mimicked by instilling inoculum into palatine tonsilar crypts. One hundred fifty days after intratonsilar inoculation, M. bovis was cultured from 30 tissues originati...
Jouglin, Maggy; Fernández-de-Mera, Isabel G; de la Cotte, Nathalie; Ruiz-Fons, Francisco; Gortázar, Christian; Moreau, Emmanuelle; Bastian, Suzanne; de la Fuente, José; Malandrin, Laurence
The diversity of Babesia species infecting cervids in parts of central and southern Spain was analyzed by collecting blood from farmed red deer (Cervus elaphus). Babesia sp. was isolated in vitro from two red deer herds in Cádiz and Ciudad Real. The number of Babesia sp. carriers differed between the two herds: 36/77 in Cádiz and 1/35 in Ciudad Real. Hyalomma lusitanicum was the most prevalent tick species identified on the Cádiz farm vegetation and on sampled animals, and is therefore a candidate vector. The molecular characteristics of 21 isolates were determined by complete (8 isolates) or partial (13 isolates) 18S rRNA gene sequencing. The sequences were highly similar (over 99.4% identity) and 6 sequence types were identified at the level of one herd only, demonstrating a rather high genetic diversity. They formed a monophyletic clade, and members of the three main sequence types shared a similar morphology and the same erythrocyte susceptibility pattern. This clade also included Babesia sp. Xinjiang isolated from sheep in China and Babesia sp. identified in giraffe in South Africa, with identities higher than 98.3% and statistically relevant phylogenetic support. None of the biological properties analyzed for both Babesia from red deer and Babesia sp. Xinjiang allowed their differentiation (ability to develop in vitro in erythrocytes from cattle and sheep, as well as in erythrocytes from different cervids, unsuccessful infection of calves). We propose the Babesia isolated from red deer as a new species named B. pecorum. Whether Babesia sp. Xinjiang and the Babesia characterized in South Africa belong to the same species is debated.
Niu, Qingli; Valentin, Charlotte; Bonsergent, Claire; Malandrin, Laurence
Rhoptry-associated-protein 1 (RAP-1) is considered as a potential vaccine candidate due to its involvement in red blood cell invasion by parasites in the genus Babesia. We examined its value as a vaccine candidate by studying RAP-1 conservation in isolates of Babesia sp. BQ1 Ningxian, Babesia sp. Tianzhu and Babesia sp. Hebei, responsible for ovine babesiosis in different regions of China. The rap-1 locus in these isolates has very similar features to those described for Babesia sp. BQ1 Lintan, another Chinese isolate also in the B. motasi-like phylogenetic group, namely the presence of three types of rap-1 genes (rap-1a, rap-1b and rap-1c), multiple conserved rap-1b copies (5) interspaced with more or less variable rap-1a copies (6), and the 3' localization of one rap-1c. The isolates Babesia sp. Tianzhu, Babesia sp. BQ1 Lintan and Ningxian were almost identical (average nucleotide identity of 99.9%) over a putative locus of about 31 Kb, including the intergenic regions. Babesia sp. Hebei showed a similar locus organization but differed in the rap-1 locus sequence, for each gene and intergenic region, with an average nucleotide identity of 78%. Our results are in agreement with 18S rDNA phylogenetic studies performed on these isolates. However, in extremely closely related isolates the rap-1 locus seems more conserved (99.9%) than the 18S rDNA (98.7%), whereas in still closely related isolates the identities are much lower (78%) compared with the 18S rDNA (97.7%). The particularities of the rap-1 locus in terms of evolution, phylogeny, diagnosis and vaccine development are discussed.
Lassen, Brian; Lepik, Triin; Bangoura, Berit
Eimeriosis is a disease that occurs globally and often affects cattle grazing on pastures contaminated with oocysts of the pathogenic species Eimeria bovis, Eimeria zuernii or Eimeria alabamensis, respectively. Nonetheless, little is understood regarding oocyst persistence on the pasture. The study was performed in the temperate climate zone. Soil samples were spiked with 100,000 E. bovis oocysts in July 2010 or with 50,000 oocysts in October 2010, respectively, both either with our without addition of cattle faeces. The soil samples were exposed to natural environmental conditions until April 2011. A subset of the samples was analysed immediately after spiking as positive control. The oocysts were recovered by a flotation method and counted in a reading chamber. On average, 23 % of the oocysts could be recovered from the positive control. The recovery of oocysts dropped to 0.30 % of the original level in the samples prepared in July independent of the addition of faeces, whereas the oocyst count was higher in the samples prepared in October, both without (2.05 %) and with (2.64 %) faecal material. No differences were observed between presence of oocysts or oocyst counts recovered in the presence or absence of faeces. Presence of faeces had a positive influence on oocyst integrity. During the winter season, the number of oocysts in the soil was lowered under the detection limit in most samples. On the other hand, the comparatively short 3-month summer period had a significant influence on the number of persisting oocysts too.
Background Bovine babesiosis is regarded as a limited health problem for Norwegian cows, and the incidence has decreased markedly since the 1930s. Rare cases of babesiosis in splenectomised humans from infection with Babesia divergens and B.venatorum have been described. The objective of this study was to determine whether birds can introduce Babesia-infected ticks. There are between 30 and 85 million passerine birds that migrate to Norway every spring. Methods Passerine birds were examined for ticks at four bird observatories along the southern Norwegian coast during the spring migrations of 2003, 2004 and 2005. The presence of Babesia was detected in the nymphs of Ixodes ricinus by real-time PCR. Positive samples were confirmed using PCR, cloning and phylogenetic analyses. Results Of 512 ticks examined, real-time PCR revealed five to be positive (1.0%). Of these, four generated products that indicated the presence of Babesia spp.; each of these were confirmed to be from Babesia venatorum (EU1). Two of the four B. venatorum-positive ticks were caught from birds having an eastern migratory route (P< 0.001). Conclusions Birds transport millions of ticks across the North Sea, the Skagerrak and the Kattegat every year. Thus, even with the low prevalence of Babesia-infected ticks, a substantial number of infected ticks will be transported into Norway each year. Therefore, there is a continuous risk for introduction of new Babesia spp. into areas where I. ricinus can survive. PMID:21699719
Oosthuizen, Marinda C; Zweygarth, Erich; Collins, Nicola E; Troskie, Milana; Penzhorn, Banie L
Babesiosis in a sable antelope (Hippotragus niger Harris, 1838) was first reported in 1930; the parasite was named Babesia irvinesmithi. Recently, specimens from an adult sable that presented with a sudden onset of disease and that subsequently died during immobilization were submitted for molecular characterization. Microscopic examination of thin blood smears revealed the presence of small piroplasms. DNA was extracted from blood samples; the V4 variable region of the 18S rRNA gene was amplified and analyzed using the reverse line blot (RLB) assay. Amplicons did not hybridize with any of the Babesia or Theileria species-specific probes present on the blot and hybridized only with a Babesia or Theileria genus-specific probe, suggesting the presence of a novel species. The full-length 18S rRNA gene sequence was obtained and aligned with published sequences of related genera, and phylogenetic trees were constructed. Sequence similarity analyses indicated that a Babesia species, designated Babesia sp. (sable), was present. The sequence showed its highest similarity to B. orientalis and to an unnamed Babesia species previously detected in bovine samples. The latter was later established to be Babesia occultans. A Babesia sp. (sable)-specific RLB oligonucleotide probe was designed and used to screen 200 South African sable samples, but so far, no other sample has been found to be positive for the presence of Babesia sp. (sable) DNA. In summary, we identified a novel piroplasm parasite from a sable antelope that died from an unknown illness. While the parasite was observed in blood smears, there is no direct evidence that it was the cause of death.
Niu, Qingli; Liu, Zhijie; Yang, Jifei; Gao, Shandian; Pan, Yuping; Guan, Guiquan; Luo, Jianxun; Yin, Hong
Babesia sp. Xinjiang is a large ovine Babesia species that was recently isolated in China. Compared with other ovine Babesia species, it has different morphological features, pathogenicity and vector tick species. The known transmitting vector is Hyalomma anatolicum. In this study, the distribution and the presence of Babesia sp. Xinjiang in small ruminants and ixodid ticks in China were assessed by specific nested-PCR assay based on the rap-1a gene. A total of 978 blood samples from sheep or goats from 15 provinces and 797 tick specimens from vegetation from 10 provinces were collected and analysed for the presence of the Babesia sp. Xinjiang. Full-length and partial rap-1a of Babesia sp. Xinjiang were amplified from field samples. The PCR results were further confirmed by DNA sequencing. Overall, 38 (3.89%) blood samples and 51 (6.4%) tick samples were positive for Babesia sp. Xinjiang infection. The highest presence (26.92%) was found in blood samples from Yunnan province, while H. qinghaiensis ticks with the highest presence of infection (21.3%) were from Gansu province. This study identified for the first time Babesia sp. Xinjiang infection in H. longicornis tick species. The rap-1a sequences of Babesia sp. Xinjiang from field blood and tick samples indicated 100% identity. The presence of Babesia sp. Xinjiang infection may increase in China. Novel potential transmitting vectors might be more extensive than previously thought.
Caimi, Karina; Romano, Maria I.; Alito, Alicia; Zumarraga, Martin; Bigi, Fabiana; Cataldi, Angel
Spoligotyping is a major tool for molecular typing of Mycobacterium bovis. This technique is based on the polymorphism of spacers that separate direct repeats (DRs) in the M. tuberculosis complex DR region. Numerous M. bovis strains show a lack of several spacers which appears as a gap in the spoligotyping pattern. To determine whether these gaps contain alternative spacers not included in the spoligotyping membrane, PCRs using primers that hybridize to the spacers adjacent to the gaps were performed. Comparing the sizes of products obtained by PCR with those deduced from spoligotyping patterns, fragments were selected and sequenced to look for alternative spacers. Upon analysis of the sequences, five alternative spacers were detected, although deletions of spacers are mainly responsible for the observed gaps. The alternative spacers, which are more frequent in M. bovis than in M. tuberculosis, may contribute to increased M. bovis differentiation. PMID:11230428
Antimicrobial treatment of persistent infection to eliminate transmission risk represents a specific challenge requiring compelling evidence of complete pathogen clearance. The limited repertoire of antimicrobial agents targeted at protozoal parasites magnifies this challenge. Using Babesia cabal...
Hanron, Amelia E; Billman, Zachary P; Seilie, Annette M; Chang, Ming; Murphy, Sean C
Babesia are increasingly appreciated as a cause of transfusion-transmitted infection. Sensitive methods are needed to screen blood products. We report herein that B. microti 18S rRNA is over 1,000-fold more abundant than its coding genes, making reverse transcription PCR (RT-PCR) much more sensitive than PCR. Babesia 18S rRNA may be useful for screening the blood supply.
Clancey, Noel; Horney, Barbara; Burton, Shelley; Birkenheuer, Adam; McBurney, Scott; Tefft, Karen
A 4-6-mo-old female red fox (Vulpes vulpes) was presented to the Atlantic Veterinary College (AVC) Teaching Hospital, Prince Edward Island, Canada. On presentation, the fox was weak and had pale mucous membranes. A complete blood count and a serum biochemistry profile were performed. Blood smear examination revealed low numbers of erythrocytes containing centrally to paracentrally located, single, rarely multiple, approximately 1 x 2 microm, oval to round organisms with morphology similar to Babesia microti. Polymerase chain reaction testing and DNA sequencing of the Babesia species 18S rRNA gene were performed on DNA extracted from whole blood. Results were positive for a Babesia microti-like parasite genetically identical to Babesia (Theileria) annae. The fox was euthanized due to poor prognosis for recovery. Necropsy examination revealed multifocal to locally extensive subacute nonsuppurative meningoencephalitis, an eosinophilic broncho-pneumonia, a moderate diffuse vacuolar hepatopathy, and lesions associated with blunt trauma to the left abdominal region. This is the first reported case of a red fox in Canada infected with a piroplasm. It remains uncertain whether the presence of this hemoparasite in this fox was pathogenic or an incidental finding. The potential for competent vectors of Babesia species on Prince Edward Island, the potential for this Babesia microti-like parasite to infect other wild and domestic canids, and the significance of this parasite to the health of infected individuals are yet to be determined.
Tadin, Ante; Turk, Nenad; Korva, Miša; Margaletić, Josip; Beck, Relja; Vucelja, Marko; Habuš, Josipa; Svoboda, Petra; Zupanc, Tatjana Avšič; Henttonen, Heikki; Markotić, Alemka
Hantaviruses, Leptospira spp., and Babesia spp. are rodent-borne pathogens present worldwide. We studied multiple co-infections of small rodents in Croatia with all three pathogens. Twenty-eight Apodemus flavicollis and 16 Myodes glareolus were tested for the presence of hantavirus RNA by real-time RT-PCR, Leptospira strains by renoculture method and Babesia DNA by PCR. Anti-hantavirus antibodies and anti-Leptospira antibodies were detected by serological methods. Very high infection rates with each pathogen were found in A. flavicollis: 20 of 28 rodents (71%) were infected with Dobrava virus, 13 rodents (46%) were infected with Leptospira, and 5 rodents (18%) were infected with Babesia. Multiple co-infections with all three pathogens were found in 3 of 28 (11%) A. flavicollis animals, suggesting that the same rodent host can be infected with several pathogens at the same time. Dual infections with both hantaviruses and Leptospira were found in 7 of 44 rodents (16%), with hantaviruses and Babesia in 2 rodents (5%), and double infection with both Leptospira and Babesia were found in 1 rodent (2%). Since hantaviruses, Leptospira, and Babesia have similar geographical distributions, it is to be expected that in other parts of the world multiple co-infections, representing a serious threat to public health, can be found.
Azmi, Kifaya; Al-Jawabreh, Amer; Nasereddin, Abedelmajeed; Abdelkader, Ahmad; Zaid, Taher; Ereqat, Suheir; Sawalha, Samer S; Baneth, Gad; Abdeen, Ziad
Dogs serve as hosts for a great number of parasites, which may affect their health and wellbeing. This study aimed to observe tick borne pathogens in dogs from Palestine including Hepatozoon canis and Babesia species. The prevalence of both H. canis and Babesia species infections in apparently healthy dogs, from ten districts of the West Bank was surveyed. DNA was extracted from blood samples obtained from dogs (n = 362) and ticks (n = 213) collected from dogs (n = 77). A primer set that amplifies a partial sequence of the Babesia and Hepatozoon 18S rRNA gene was used for PCR and the DNA sequences of the PCR products of all samples were determined. Twenty-nine (8·0%) of the dogs were found infected including 20 with H. canis (5·5%), seven with Babesia vogeli (1·9%) and two with undefined Babesia spp. (0·6%). Twelve Rhipicephalus sanguineus s.l ticks were pathogen-positive, including ten with H. canis (4·7%), one with B. vogeli (0·5%), and one with Hepatozoon felis (0·5%). The results indicated that a wide range of tick borne pathogens is circulating in the canine population in the surveyed region. This study is the first report on the prevalence of H. canis, B. vogeli and Babesia spp. in dogs in Palestine and its results will assist in the management of diseases associated with these blood parasites.
Williams, Brianna M; Berentsen, Are; Shock, Barbara C; Teixiera, Maria; Dunbar, Michael R; Becker, Matthew S; Yabsley, Michael J
A molecular survey was conducted for several hemoparasites of domestic dogs and three species of wild carnivores from two sites in Zambia. Three Babesia spp. were detected including Babesia felis and Babesia leo in lions (Panthera leo) and a Babesia sp. (similar to Babesia lengau) in spotted hyenas (Crocuta crocuta) and a single lion. All wild dogs (Lycaon pictus) and domestic dogs were negative for Babesia. High prevalences for Hepatozoon were noted in all three wild carnivores (38-61%) and in domestic dogs (13%). Significantly higher prevalences were noted in hyenas and wild dogs compared with domestic dogs and lions. All carnivores were PCR negative for Ehrlichia canis, Ehrlichia ewingii, and Bartonella spp. Overall, high prevalences and diversity of Babesia and Hepatozoon were noted in wild carnivores from Zambia. This study is the first molecular characterization of Babesia from any hyena species and is the first report of a Babesia sp. closely related to B. lengau, a parasite previously only reported from cheetahs (Acinonyx jubatus), in lions and hyenas. Although usually benign in wild carnivores, these hemoparasites can be pathogenic under certain circumstances. Importantly, data on vectors for these parasites are lacking, so studies are needed to identify vectors as well as determine transmission routes, infection dynamics, and host specificity of these hemoparasites in wildlife in Africa and also the risk of transmission between domestic animals and wildlife.
Dedet, J P; Veilly, M; Robin, Y; Bonnevie, O; Landau, I
A two-toed sloth (Choloepus didactylus) from French Guiana showed an erythrocytic parasite morphologically related to the Babesiidae family, and described as Babesia choloepi. It is the first Babesia species described in the Edentata order.
Cattle were inoculated with Mycobacterium bovis, Mycobacterium tuberculosis, or Mycobacterium kansasii to compare antigen-specific immune responses to varied patterns of mycobacterial disease. Disease expression ranged from colonization with associated pathology (M. bovis), colonization without path...
Brasseur, P; Lecoublet, S; Kapel, N; Favennec, L; Ballet, J J
The susceptibilities of three bovine and two human Babesia divergens isolates to antimicrobial agents were evaluated in vitro by a tritiated hypoxanthine incorporation assay. The MICs at which 50% of isolates are inhibited (MIC50s) for mefloquine (chlorhydrate), chloroquine (sulfate), quinine (chlorhydrate), clindamycin (phosphate), pentamidine (isethionate), phenamidine (isethionate) plus oxomemazine (chlorhydrate), lincomycin (chlorhydrate monohydrate), and imidocarb (dipropionate) were determined. Except for imidocarb, the MIC50s observed for the different isolates were close. Imidocarb and the combination of phenamidine plus oxomemazine exhibited the highest in vitro activity, while antimalarial agents such as mefloquine, choroquine, and quinine were inactive. Other drugs had intermediate activities. The data support further in vitro evaluation of antimicrobial agents active against B. divergens for the improvement of therapeutic strategies.
Brasseur, Philippe; Lecoublet, Sophie; Kapel, Nathalie; Favennec, Loic; Ballet, Jean J.
The susceptibilities of three bovine and two human Babesia divergens isolates to antimicrobial agents were evaluated in vitro by a tritiated hypoxanthine incorporation assay. The MICs at which 50% of isolates are inhibited (MIC50s) for mefloquine (chlorhydrate), chloroquine (sulfate), quinine (chlorhydrate), clindamycin (phosphate), pentamidine (isethionate), phenamidine (isethionate) plus oxomemazine (chlorhydrate), lincomycin (chlorhydrate monohydrate), and imidocarb (dipropionate) were determined. Except for imidocarb, the MIC50s observed for the different isolates were close. Imidocarb and the combination of phenamidine plus oxomemazine exhibited the highest in vitro activity, while antimalarial agents such as mefloquine, choroquine, and quinine were inactive. Other drugs had intermediate activities. The data support further in vitro evaluation of antimicrobial agents active against B. divergens for the improvement of therapeutic strategies. PMID:9559789
Mycoplasma bovis (M. bovis) has recently emerged as a significant and costly infectious disease problem in bison, generally presenting as severe, caseonecrotic pneumonia. Here we describe three diagnostic cases in which M. bovis is strongly implicated as a causative agent of necrotic pharyngitis. ...
... 9 Animals and Animal Products 2 2011-01-01 2011-01-01 false Tapeworm cysts (cysticercus bovis) in... PARTS § 311.23 Tapeworm cysts (cysticercus bovis) in cattle. (a) Except as provided in paragraph (b) of... humerus. (2) Carcasses of cattle showing one or more tapeworm lesions of cysticercus bovis but not...
... 9 Animals and Animal Products 2 2010-01-01 2010-01-01 false Tapeworm cysts (cysticercus bovis) in... PARTS § 311.23 Tapeworm cysts (cysticercus bovis) in cattle. (a) Except as provided in paragraph (b) of... humerus. (2) Carcasses of cattle showing one or more tapeworm lesions of cysticercus bovis but not...
... 9 Animals and Animal Products 2 2014-01-01 2014-01-01 false Tapeworm cysts (cysticercus bovis) in... PARTS § 311.23 Tapeworm cysts (cysticercus bovis) in cattle. (a) Except as provided in paragraph (b) of... humerus. (2) Carcasses of cattle showing one or more tapeworm lesions of cysticercus bovis but not...
... 9 Animals and Animal Products 2 2012-01-01 2012-01-01 false Tapeworm cysts (cysticercus bovis) in... PARTS § 311.23 Tapeworm cysts (cysticercus bovis) in cattle. (a) Except as provided in paragraph (b) of... humerus. (2) Carcasses of cattle showing one or more tapeworm lesions of cysticercus bovis but not...
... 9 Animals and Animal Products 2 2013-01-01 2013-01-01 false Tapeworm cysts (cysticercus bovis) in... PARTS § 311.23 Tapeworm cysts (cysticercus bovis) in cattle. (a) Except as provided in paragraph (b) of... humerus. (2) Carcasses of cattle showing one or more tapeworm lesions of cysticercus bovis but not...
Mycobacterium bovis infection of cats is exceedingly rare in non-endemic regions for bovine tuberculosis. This case study describes the diagnosis and clinical management of pulmonary M. bovis infection in two indoor-housed cats and their association with at least one M. bovis-infected human in Texas...
Stalheim, O H; Proctor, S J
Two pregnant cows aborted 11 and 18 days after Mycoplasma agalactiae subsp bovis was inoculated into the amniotic fluids. The placentas were retained. The fetuses (approx 100 and 150 days of age) were decomposed; M agalactiae subsp bovis was recovered from several tissues of the fetuses, the placentas, and fetal fluids. The same organism was given by intraperitoneal injection to 2 other pregnant (130 and 180 days, respectively) cows. At necropsy of the latter 36 days later, placentitis was severe; M agalactiae subsp bovis was recovered from the placentas of both cows and from the fetus of 1 cow. Control cows given sterile mycoplasma cultural medium by intraamnion or intraperitoneal injection did not abort and were not infected. When first recovered from the bovine placenta and fetus, M agalactiae subsp bovis grew slowly in liquid medium and assumed bizarre colonial morphology on solidified medium. Colonies were small (0.1 to 0.5 mm) and dark and lacked halos, but they reacted specifically in the direct fluorescent antibody test with equine M agalactiae subsp bovis antiserum. After 1 or 2 subcultures, the isolates grew at a normal rate and displayed their usual colonial morphology.
Irvin, R T; To, M; Costerton, J W
Alysiella bovis adheres to surfaces by means of short, ruthenium red-staining, rod-like fimbriae. The fimbriae remain associated with the cell envelope of A. bovis, even when sonicated or exposed sequentially to toluene, Triton X-100, lysozyme, ribonuclease, and deoxyribonuclease. Adhesion of outer membrane-derived cell wall ghosts of A. bovis to glass was inhibited by IO4-, sodium dodecyl sulfate, urea, pronase, and trypsin. Protease treatment digested the fimbriae from the distal end, and exposure to sodium dodecyl sulfate depolymerized the fimbriae. Exposure of ghosts to 1% sodium dodecyl sulfate preferentially solubilized a 16,500-dalton protein which was subsequently purified by gel filtration and demonstrated to be a glycoprotein (ca. 17% carbohydrate). Antibodies raised against the 16,500-dalton glycoprotein agglutinated whole cells and inhibited adhesion of ghosts to glass. Images PMID:6209260
Faurholt-Jepsen, Daniel; Lillebaek, Troels; Nielsen, Ming-Yuan; Nielsen, Susanne Dam
In Denmark, tuberculous meningitis is rare. Central nervous system (CNS) involvement with Mycobacterium bovis is even rarer and has only been seen three times since 1992. We present a case of M. bovis meningitis in a previously healthy young Nigerian-born male, who had been exposed to unpasteurized dairy products in Nigeria but had no known contact with larger mammals. Before the development of meningitis, the patient had several contacts with the health system due to fever and non-specific symptoms. Finally, upon hospital admission, the patient was diagnosed with M. tuberculosis complex meningitis and treated empirically. After 13 days he was discharged without neurological sequelae. Later, the culture revealed M. bovis and treatment was adjusted accordingly.
Vazquez-Chacon, Carlos A; Martínez-Guarneros, Armando; Couvin, David; González-Y-Merchand, Jorge A; Rivera-Gutierrez, Sandra; Escobar-Gutierrez, Alejandro; De-la-Cruz López, Juan J; Gomez-Bustamante, Adriana; Gonzalez-Macal, Gabriela A; Gonçalves Rossi, Livia Maria; Muñiz-Salazar, Raquel; Rastogi, Nalin; Vaughan, Gilberto
Here, we describe the molecular characterization of six human Mycobacterium bovis clinical isolates, including three multidrug resistant (MDR) strains, collected in Mexico through the National Survey on Tuberculosis Drug Resistance (ENTB-2008), a nationally representative survey conducted during 2008-2009 in nine states with a stratified cluster sampling design. The genetic background of bovine M. bovis strains identified in three different states of Mexico was studied in parallel to assess molecular relatedness of bovine and human strains. Additionally, resistance to first and second line anti-tuberculosis (TB) drugs and molecular identification of mutations conferring drug resistance was also performed. All strains were characterized by spoligotyping and 24-loci MIRU-VNTRs, and analyzed using the SITVIT2 (n = 112,000 strains) and SITVITBovis (n = 25,000 strains) proprietary databases of Institut Pasteur de la Guadeloupe. Furthermore, data from this study (n = 55 isolates), were also compared with genotypes recorded for M. bovis from USA (n = 203), Argentina (n = 726), as well as other isolates from Mexico (independent from the present study; n = 147), to determine any evidence for genetic relatedness between circulating M. bovis strains. The results showed that all human M. bovis cases were not genetically related between them or to any bovine strain. Interestingly, a high degree of genetic variability was observed among bovine strains. Several autochthonous and presumably imported strains were identified. The emergence of drug-resistant M. bovis is an important public health problem that jeopardizes the success of TB control programs in the region.
Galdy, Salvatore; Nastasi, Giuseppe
A relationship between infective endocarditis and colon cancer was established in 1950, and Streptococcus bovis was successfully isolated in 1970. However, this association and its pathogenesis still remain unclear. In this paper, we describe the clinical case of a patient with a history of colon cancer and infective endocarditis caused by Streptococcus bovis. The role of S bovis as an aetiological agent in the development of colon cancer is intriguing but uncertain. S bovis infection should be considered a silent sign of gastrointestinal malignancy or hepatic disease. We believe that in order to demonstrate the presence of colon cancer, all patients with S bovis infection require an endoscopic investigation of the colon.
Gil-Turnes, C; Ribeiro, G A
Several properties of the adhesins of eight isolates of Moraxella bovis recovered from cattle suffering from infectious keratoconjunctivitis, were studied. Adhesions were detected through autoagglutination in saline and hemagglutination. Autoagglutinating strains agglutinated red blood cells of the chicken, rabbit, sheep and swine, but not those of the guinea pig. The adhesins were not inhibited by D-mannose or D-galactose and resisted heating at 100 degrees C for 15 minutes. Magnesium chloride at a final concentration of 10% inhibited autoagglutination and hemagglutination. The value of the hemagglutination test for monitoring synthesis of fimbriae by M. bovis, is discussed. PMID:3986674
Birkenheuer, Adam J; Levy, Michael G; Stebbins, Martha; Poore, Matthew; Breitschwerdt, Edward
Stray dogs (n=359) and kennel dogs (n=149) from North Carolina were tested for evidence of antiBabesia antibodies. AntiBabesia antibodies were detected in 21/359 and 22/149 of the stray and kennel dogs, respectively. A total of 57 dogs from both groups were tested for babesiasis by light microscopy and polymerase chain reaction (PCR). Babesia deoxyribonucleic acid (DNA) was detected in 3/28 of the stray dogs and 14/29 of the kennel dogs. When Babesia DNA was detected by PCR, the species-specific PCR results differed from the Babesia species antibody titer results in 6/17 of the PCR-positive dogs. There was no association between antiBabesia antibodies and the presence of ticks. There are currently Babesia gibsoni epizootics affecting American pit bull terrier kennels.
Leclaire, Sarah; Menard, Sandie; Berry, Antoine
Piroplasms, including Babesia, Cytauxzoon and Theileria species, frequently infect domestic and wild mammals. At present, there is no information on the occurrence and molecular identity of these tick-borne blood parasites in the meerkat, one of South Africa's most endearing wildlife celebrities. Meerkats live in territorial groups, which may occur on ranchland in close proximity to humans, pets and livestock. Blood collected from 46 healthy meerkats living in the South-African Kalahari desert was screened by microscopy and molecular methods, using PCR and DNA sequencing of 18S rRNA and ITS1 genes. We found that meerkats were infected by 2 species: one species related to Babesia sp. and one species related to Cytauxzoon sp. Ninety one percent of the meerkats were infected by the Cytauxzoon and/or the Babesia species. Co-infection occurred in 46% of meerkats. The pathogenicity and vectors of these two piroplasm species remains to be determined.
Drewe, J A; Foote, A K; Sutcliffe, R L; Pearce, G P
Pathological lesions associated with Mycobacterium bovis infection (bovine tuberculosis; bTB) in free-living meerkats (Suricata suricatta) in the Kalahari Desert of South Africa are described. The pathology of bTB in meerkats was determined through detailed post-mortem examinations of 57 animals (52 meerkats showing clinical signs of bTB, and five not showing signs of disease). Lymph nodes and tissue lesions thought to be associated with bTB were cultured for mycobacteria. All 52 bTB-infected meerkats showed gross or microscopical granulomatous lesions, but M. bovis was cultured from only 42% (22/52) of these animals. The majority (96%, 50/52) of diseased meerkats had lesions in multiple sites, the pattern of which suggested haematogenous spread of M. bovis infection in this species. The histological characteristics of the tuberculous lesions, together with the gross pathology and the wide range of body systems affected, indicate that infection in meerkats is acquired principally via the respiratory and oral routes, whereas excretion is most likely via the respiratory tract and suppurating skin wounds. Urine and faeces appear to be unlikely sources of infection. The findings of this study provide information on the transmission, pathogenesis and epidemiology of bTB in meerkats that is likely to be relevant to the understanding of M. bovis infection in other social mammal species such as the European badger (Meles meles).
Three isolates of Moraxella bovis, recovered from cattle with signs of infectious bovine keratoconjunctivitis, were tested for autoagglutinating activity, hemagglutinating activity and pathogenicity in young calves. Only the autoagglutinating and hemagglutinating isolates were pathogenic in calves. Treatment of the pathogenic isolates with magnesium chloride eliminated their pathogenic effects. PMID:6667436
... samples of completed product from each serial shall be tested as follows: (1) Protein concentration. The final product shall contain a protein concentration of 1.0 ±0.1 mg/ml. The Microkjeldahl Test for..., Intradermic is a purified protein derivative produced from cultures of Mycobacterium bovis Strain...
... samples of completed product from each serial shall be tested as follows: (1) Protein concentration. The final product shall contain a protein concentration of 1.0 ±0.1 mg/ml. The Microkjeldahl Test for..., Intradermic is a purified protein derivative produced from cultures of Mycobacterium bovis Strain...
Introduction. Cattle infected with Mycobacterium bovis, the causative agent of bovine tuberculosis and a relevant zoonosis to humans, may be sent to slaughter before diagnosis of infection because of slow multiplication of the pathogen. Purpose. This study evaluates multiple processing interventi...
Behrendt, Jan Hillern; Hermosilla, Carlos; Hardt, Martin; Failing, Klaus; Zahner, Horst; Taubert, Anja
For successful in vivo infection, Eimeria bovis sporozoites have to traverse the mucosal layer of the ileum to infect lymphatic endothelial cells and may, thereby, be exposed to the interstitial fluid and to the lymph representing potential targets for leukocytes. To mimic this situation in vitro, we exposed E. bovis sporozoites to bovine PMN and found enhanced elimination of the parasites. Addition of immune serum clearly increased these reactions, whereas neonatal calf serum had no effect, thus proposing a PMN-derived antibody-dependent cytotoxicity. Scanning and transmission electron microscopy showed PMN engulfing sporozoites or extending filopodia towards them and occasionally incorporating the parasites. PMN reacted with enhanced transcription of IL-6, MCP-1, GROalpha, TNF-alpha, and iNOS genes after exposure to sporozoites while stimulation with merozoite-antigen, in addition, upregulated IL-8, IP-10 and IL-12 gene transcription. Furthermore, enhanced in vitro oxidative burst and phagocytic activities were observed after contact of PMN with viable sporozoites. To verify the potential role of PMN in the in vivo situation, we analysed the general phagocytic and oxidative burst activities of PMN obtained ex vivo from E. bovis experimentally infected calves. Enhanced levels of both activities were found early p.i. (1-5 days) and towards the end of the first schizogony (days 13-22 p.i.) underlining the in vitro data. Our results suggest that PMN-mediated, innate immune reactions play an important role in the early immune response to E. bovis infections in calves.
... Tuberculin supplied by Animal and Plant Health Inspection Service. (1) Test animals. White female guinea pigs... used in a previous test, shall be used in the specificity test. Twenty-three guinea pigs (10 sensitized... being tested, and 20 guinea pigs (10 sensitized with M. bovis and 10 sensitized with M. avium) shall...
Ramarokoto, H; Andrianasolo, D; Rasolonavalona, T; Ramaroson, F; Razafitsiarovana, I; Vincent, V; Ratsimba, L; Rasolofo Razanamparany, V
We report a chronic case of pulmonary tuberculosis in a Malagasy citizen from Antsohihy (West of Madagascar), who was infected with a multi-drug resistant Mycobacterium bovis strain. This is the first case reported of the isolation of such a strain in Madagascar.
Fathi, Yashar; Golmaii, Poone; Rabiee, Atoosa; Samadpoor, Ali; Shahverdi, Nooshin; Nikbin, Behrooz
The effects of laser on the immune system have not been extensively characterized. Low power laser sources, such as GaAlAs laser have been found to produce photo biological effects with evidence of interference with immunological functions. We have investigated the effects of GaAlAs laser irradiation on the immune response due to Mycobacterium bovis BCG infection in mice. BALB/C mice were exposed on the abdomen skin to GaAlAs laser radiation (810 nm) for 3 consequent days (Days = -2, -1, 0) before infection with 1 x 106 live units of Mycobacterium bovis Bacillus Calmette-Guerin (BCG) in the footpad. 21 days later groups of mice were tested for a delayed type hypersensitivity (DTH) response to the purified protein derivative (PPD) of tubercle bacilli and the course of infection was monitored by measuring the size of the infected footpad. In the mice treated by laser, the DTH response to PPD was significantly suppressed (P-value<0.045) compared to unirradiated mice, when tested 21 days after BCG infection. When laser irradiated 13 days after BCG infection (Days: +13, +14, +15) BALB/C mice did not show a significant decrease in their DTH response to PPD indicating that the laser-induced suppression of BCG occurs only at the induction stage of the immune response. Thus mice exposed to the laser radiation before BCG infection showed an impaired DTH response to Mycobacterium, whereas mice exposed to the laser irradiation after BCG did not. These studies demonstrate that a systemic effect of laser irradiation can suppress the development and expression of immunity to pathogenic bacteria in mice. This suppression could be at the induction stage of the immune response (DTH) but not the elicitation stage. Also it seems that laser radiation, potentially, could have side effects for the immune system, on the basis of suppression effects it has shown on DTH response.
Usatii, Natalia; Khachatrian, Aelita; Stratidis, John
This article describes the case of spontaneous splenic rupture as a rare complication of infection with Babesia species. We will discuss the symptomatology that this disease could present along with both surgical and non-surgical management approaches. Babesia infection often presents with mild to moderate symptoms, but can rapidly progress to significant injury including splenic rupture. The first case reported in a medical journal was in 2007. Treatment usually involves a two-drug regimen; clindamycin plus quinine, or atovaquone plus azithromycin (as in our patient). If hemodynamic stability is present, a primary non-surgical treatment may be especially beneficial since splenectomy may worsen optimal immunologic function and the infection itself.
Iguchi, Aiko; Matsuu, Aya; Matsuyama, Kazuyoshi; Hikasa, Yoshiaki
Artemisinin has many derivatives, and it is effective against Plasmodium spp. However, only a limited number of reports have confirmed the efficacy of artemisinin derivatives against Babesia spp. In this study, whether artemisinin and artemether could inhibit the growth of Babesia gibsoni was evaluated in vitro. In addition, the interaction between artemether and lumefantrine was evaluated. These drugs inhibited the growth of B. gibsoni, but artemisinin and artemether showed lower sensitivity against atovaquone-resistant B. gibsoni than against wild-type B. gibsoni. The interaction between artemether and lumefantrine showed synergism against B. gibsoni. Although further study is needed, the combination of artemisinin derivatives could be useful for babesiosis.
Jia, H; Terkawi, M A; Aboge, G O; Goo, Y-K; Luo, Y; Li, Y; Yamagishi, J; Nishikawa, Y; Igarashi, I; Sugimoto, C; Fujisaki, K; Xuan, X
Peptidases of parasitic protozoa are currently under intense investigation in order to identify novel virulence factors, drug targets, and vaccine candidates, except in Babesia. Leucine aminopeptidases in protozoa, such as Plasmodium and Leishmania, have been identified to be involved in free amino acid regulation. We report here the molecular and enzymatic characterization, as well as the localization of a leucine aminopeptidase, a member of the M17 cytosolic aminopeptidase family, from B. gibsoni (BgLAP). A functional recombinant BgLAP (rBgLAP) expressed in Escherichia coli efficiently hydrolysed synthetic substrates for aminopeptidase, a leucine substrate. Enzyme activity of the rBgLAP was found to be optimum at pH 8.0 and at 37 degrees C. The substrate profile was slightly different from its homologue in P. falciprum. The activity was also strongly dependent on metal divalent cations, and was inhibited by bestatin, which is a specific inhibitor for metalloprotease. These results indicated that BgLAP played an important role in free amino acid regulation.
González, Luis M; Castro, Emma; Lobo, Cheryl A; Richart, Alberto; Ramiro, Raquel; González-Camacho, Fernando; Luque, Daniel; Velasco, Aurelio C; Montero, Estrella
Human babesiosis is a zoonosis primarily transmitted through Ixodes ticks and alternatively by routes such as blood transfusions from asymptomatic donors. We report the first case of human babesiosis caused by Babesia divergens in a patient with HIV. This study also focuses on elucidating the possible transmission route of infection in this patient, who received numerous blood transfusions but showed patent symptoms only after splenectomy. A battery of detection tools along with a novel Western-Blot Assay and Enzyme Linked Immunosorbent Assay using the major surface protein of B. divergens (Bd37) as a target were used to evaluate the presence of B. divergens or antibodies against the parasite in samples from the patient and the blood donors involved in this case. A retrospective study of the humoral status against the parasite revealed B. divergens IgG antibodies in one of the implicated donors, but also showed that the patient had been already exposed to the parasite before any transfusion. Thus, this analysis of natural and transfusion transmission routes suggests a pre-existing subclinical babesiosis in the patient.
Shimada, Kayoko; Azuma, Yuko; Kawase, Masaya; Takahashi, Toshiharu; Schaffer, Stephen W; Takahashi, Kyoko
Calculus Bovis (C. Bovis) is a commonly used animal-derived therapeutic preparation. To meet the increasing clinical demand for the preparation, two artificial substitutes for Bos Taurus have been introduced in China: artificial C. Bovis and in vitro cultured C. Bovis. However, information on their efficacy and safety is inadequate. Therefore, we investigated the biological differences between the commonly used natural preparation and its two substitutes, with the aim of not only identifying the differences but also providing a procedure to distinguish between the different preparations.In the study, we prepared 9 natural C. Bovis, 2 artificial C. Bovis, and 2 in vitro cultured C. Bovis preparations for evaluation. Differences were noted between the three preparations relative to their effect on viability of cardiac fibroblasts from 1-day-old Wistar rats. Although natural C. Bovis had no effect on cell viability, 1-h treatment of the cells with 0.25 mg/ml of the substitutes significantly reduced cell viability, as detected by the MTS assay. Based on liquid chromatography and inductively coupled plasma mass spectrometry, the preparations also differed in composition. Indeed, the substitutes contained more taurine, cholic acid, iron, magnesium, and calcium than the natural preparations. They also differed spectroscopically.The present results reveal significant biological differences between natural C. Bovis and two of its substitutes. Since the substitutes appear to contain more taurine, cholic acid, and elements, these constituents may serve as markers to distinguish between natural C. Bovis and its substitutes.
Burr, Holly N; Wolf, Felix R; Lipman, Neil S
Corynebacterium bovis is a common pathogen in athymic nude mouse colonies. Control and eradication of the organism are challenging because depopulation and restricted colony access are often not options within vivaria. We evaluated potential sources and dissemination routes of C. bovis in an enzootically infected colony. Immunocompetent mice and personnel were evaluated for their potential to carry C. bovis, and husbandry and sanitation methods were evaluated for their efficacy in preventing cross-contamination. C. bovis was detected in furred immunocompetent mice previously exposed to infected athymic nude mice and in the nasopharynx of humans. Microisolation cages were not effective in maintaining athymic nude mice C. bovis-free when they were housed in a room known to contain immunodeficient mice with C. bovis infections. A tunnel washer that provided a ≥180 °F final rinse provided effective elimination of C. bovis from cage components. Passive and active air sampling techniques showed airborne dispersal of C. bovis despite the use of individually ventilated caging systems and stringent operational standards. Bacterial growth was not observed in settle plates placed inside autoclaved individually ventilated microisolation cages on various ventilated racks for 24-h periods. C. bovis aerosolization was shown to be a means of spread of the bacterium during cage-change procedures inside a class II type A2 biosafety cabinet. Our findings indicate that C. bovis can be a pervasive environmental contaminant in infected rodent holding rooms and successful eradication strategies must include environmental decontamination and attention to air quality.
Battsetseg, B; Matsuo, T; Xuan, X; Boldbaatar, D; Chee, S H; Umemiya, R; Sakaguchi, T; Hatta, T; Zhou, J; Verdida, A R; Taylor, D; Fujisaki, K
Ornithodoros moubata ticks were fed on blood infected with Babesia equi. However, the parasites were quickly cleared as evidenced by the disappearance of B. equi-specific ribosomal RNA from the ticks. We hypothesized that if the Babesia parasite can escape midgut-associated barriers a non-vector tick can become infected with Babesia. To test this hypothesis, B. equi parasite-infected blood from in vitro culture was injected into the haemocoel of ticks. B. equi-specific rRNA was surprisingly detected 45 days after injection even in the eggs. Babesia-free dogs were infested with O. moubata ticks that were infected by inoculation with B. gibsoni-infected red blood cells. Parasitaemia and antibody production against Bg-TRAP of B. gibsoni increased gradually. These results indicate that O. moubata may be a useful vector model for Babesia parasites and also a very important tool for studies on tick immunity against Babesia parasites and tick-Babesia interactions.
Wildlife reservoirs of Mycobacterium bovis represent serious obstacles to the eradication of tuberculosis in domestic livestock. The cause for many faltering eradication programs is the presence of wildlife reservoirs of M. bovis. One approach in dealing with this wildlife reservoir is to vaccinate ...
White-tailed deer (Odocoileus virginianus) serve as a reservoir for bovine tuberculosis, caused by Mycobacterium bovis, and can be a source of infection in cattle. Vaccination with M. bovis bacille Calmette-Guerin (BCG) is being considered for management of bovine tuberculosis in deer. Presently, no...
Rizzi, Caroline; Peiter, Ana Carolina; Oliveira, Thaís Larré; Seixas, Amilton Clair Pinto; Leal, Karen Silva; Hartwig, Daiane Drawanz; Seixas, Fabiana Kommling; Borsuk, Sibele; Dellagostin, Odir Antônio
BACKGROUND Bovine tuberculosis (TB) is a zoonotic disease caused by Mycobacterium bovis, responsible for causing major losses in livestock. A cost effective alternative to control the disease could be herd vaccination. The bacillus Calmette-Guérin (BCG) vaccine has a limited efficacy against bovine TB, but can improved by over-expression of protective antigens. The M. bovis antigen 85B demonstrates ability to induce protective immune response against bovine TB in animal models. However, current systems for the construction of recombinant BCG expressing multiple copies of the gene result in strains of low genetic stability that rapidly lose the plasmid in vivo. Employing antibiotic resistance as selective markers, these systems also compromise vaccine safety. We previously reported the construction of a stable BCG expression system using auxotrophic complementation as a selectable marker. OBJECTIVES The fundamental aim of this study was to construct strains of M. bovis BCG Pasteur and the auxotrophic M. bovis BCG ΔleuD expressing Ag85B and determine their stability in vivo. METHODS Employing the auxotrophic system, we constructed rBCG strains that expressed M. bovis Ag85B and compared their stability with a conventional BCG strain in mice. Stability was measured in terms of bacterial growth on the selective medium and retention of antigen expression. FINDINGS The auxotrophic complementation system was highly stable after 18 weeks, even during in vivo growth, as the selective pressure and expression of antigen were maintained comparing to the conventional vector. MAIN CONCLUSION The Ag85B continuous expression within the host may generate a stronger and long-lasting immune response compared to conventional systems. PMID:28177046
Vargas-Hernández, G; André, M R; Faria, J L M; Munhoz, T D; Hernandez-Rodriguez, M; Machado, R Z; Tinucci-Costa, M
Ehrlichiosis and babesiosis are tick-borne diseases, caused mainly by Ehrlichia canis and Babesia canis, respectively, with a worldwide occurrence in dogs, whose main vector is the brown-dog tick, Rhipicephalus sanguineus. The present work aimed to detect the presence of E. canis and Babesia sp. in 91 dog blood samples in Colombia, by molecular and serological techniques. We also performed sequence alignment to indicate the identity of the parasite species infecting these animals. The present work shows the first molecular detection of E. canis and B. vogeli in dogs from Colombia. Immunoglobulin-G (IgG) antibodies to E. canis and Babesia vogeli were found in 75 (82.4%) and 47 (51.6%) sampled dogs, respectively. Thirty-seven (40.6%) and 5 (5.5%) dogs were positive in PCR for E. canis and Babesia sp., respectively. After sequencing, amplicons showed 99% of identity with isolates of E. canis and B. vogeli. The phylogenetic trees based on 16S rRNA-Anaplasmataceae sequences and 18S rRNA-piroplasmid sequences supported the identity of the found E. canis and B. vogeli DNAs, respectively. The present work shows the first molecular detection of E. canis and B. vogeli in dogs in Colombia.
Penzhorn, B L; Lewis, B D; López-Rebollar, L M; Swan, G E
The efficacy of 5 drugs was tested against experimental Babesia felis infection in domestic cats. Two of the drugs, rifampicin and a sulphadiazine-trimethoprim combination, appeared to have an anti-parasitic effect, but were inferior to primaquine. The other 3 drugs, buparvaquone, enrofloxacin and danofloxacin, had no significant anti-babesial effect.
Vanstreels, Ralph Eric Thijl; Woehler, Eric J.; Ruoppolo, Valeria; Vertigan, Peter; Carlile, Nicholas; Priddel, David; Finger, Annett; Dann, Peter; Herrin, Kimberly Vinette; Thompson, Paul; Ferreira Junior, Francisco C.; Braga, Érika M.; Hurtado, Renata; Epiphanio, Sabrina; Catão-Dias, José Luiz
Blood parasites are potential threats to the health of penguins and to their conservation and management. Little penguins Eudyptula minor are native to Australia and New Zealand, and are susceptible to piroplasmids (Babesia), hemosporidians (Haemoproteus, Leucocytozoon, Plasmodium) and kinetoplastids (Trypanosoma). We studied a total of 263 wild little penguins at 20 sites along the Australian southeastern coast, in addition to 16 captive-bred little penguins. Babesia sp. was identified in seven wild little penguins, with positive individuals recorded in New South Wales, Victoria and Tasmania. True prevalence was estimated between 3.4% and 4.5%. Only round forms of the parasite were observed, and gene sequencing confirmed the identity of the parasite and demonstrated it is closely related to Babesia poelea from boobies (Sula spp.) and B. uriae from murres (Uria aalge). None of the Babesia-positive penguins presented signs of disease, confirming earlier suggestions that chronic infections by these parasites are not substantially problematic to otherwise healthy little penguins. We searched also for kinetoplastids, and despite targeted sampling of little penguins near the location where Trypanosoma eudyptulae was originally reported, this parasite was not detected. PMID:25853053
A novel large Babesia sp. from an infected dog was cultivated in vitro by microaerophilous stationary phase culture methodology. uRPMI-1640 medium supplemented with 40% canine serum, Albumax, hypoxanthine and thymidine with antibiotics and buffered with HEPES supported the primary culture of the pa...
Spada, Eva; Proverbio, Daniela; Galluzzo, Paola; Perego, Roberta; Roggero, Nora; Caracappa, Santo
Emerging diseases caused by piroplasms pose a health risk for man and other animals, and domestic cats have been proposed as potential reservoirs for some piroplasm infections. The aim of this study was to identify the frequency of the piroplasms Babesia microti and Cytauxzoon felis in stray cats from northern Italy and to identify possible risk factors associated with these infections. Blood samples from 260 stray cats enrolled in a trap-neuter-release (TNR) program in northern Italy were examined with conventional PCR for the presence of Babesia microti and Cytauxzoon felis DNA. No sample (0.0%) tested positive for C. felis, whilst B. microti DNA was detected in two samples (0.8%). Both infected cats were in good clinical condition and recovered well from the neutering surgery. One of these two cats had a triple coinfection with Babesia microti, Candidatus Mycoplasma haemominutum, and Anaplasma phagocytophilum. Evidence presented in this study indicates that the blood borne protozoans Babesia microti and Cytauxzoon felis are not widely distributed in stray cat populations in Milan, northern Italy, and that the significance of cats as a reservoir host for B. microti in this area is limited. PMID:24895629
Sivakumar, Thillaiampalam; Tattiyapong, Muncharee; Okubo, Kazuhiro; Suganuma, Keisuke; Hayashida, Kyoko; Igarashi, Ikuo; Zakimi, Satoshi; Matsumoto, Kotaro; Inokuma, Hisashi; Yokoyama, Naoaki
Babesia ovata is a tick-transmitted hemoprotozoan parasite of cattle. In the present study, we analyzed tick DNA samples (n=1459) prepared from questing ticks collected from various cattle pastures in Hokkaido (Shibecha, Taiki, Otofuke, Memuro, and Shin-Hidaka districts) and Okinawa (Yonaguni Island) prefectures of Japan for B. ovata. When all the tick DNA samples were screened by a previously described B. ovata-specific apical membrane antigen-1 (AMA-1) gene-based polymerase chain reaction (PCR) assay, none of the DNA samples was positive. Therefore, we developed a PCR assay based on the protozoan beta-tubulin (β-tubulin) gene to detect B. ovata from ticks in Japan. In the specificity test, the PCR assay amplified the expected 444-bp target gene fragment from B. ovata DNA. No PCR products were amplified from DNA samples from other blood pathogens, bovine blood, or ticks. In addition, the PCR assay detected 100 fg of B. ovata-genomic DNA extracted from an in vitro culture of the parasites. Subsequently, when all the tick DNA samples were screened by this new PCR assay, 18 were positive for B. ovata. Positive samples were found only in the Yonaguni and Memuro areas. In Okinawa, where all the ticks were identified as Haemaphysalis longicornis, 9.7% of the samples were PCR-positive, while a single tick (Ixodes ovatus) from Memuro was infected with B. ovata. When the nucleotide sequences of the PCR amplicons were phylogenetically analyzed, they formed a separate clade containing a previously described β-tubulin gene sequence from B. ovata (Miyake strain), confirming that the PCR assay had detected only B. ovata from the tick DNA samples. This is the first report that describes the PCR detection of B. ovata in ticks. The findings warrant transmission experiments to evaluate I. ovatus as a potential vector of B. ovata.
Spositto, F L E; Campanerut, P A Z; Ghiraldi, L D; Leite, C Q F; Hirata, M H; Hirata, R D C; Siqueira, V L D; Cardoso, R Fressatti
We evaluated a multiplex-PCR to differentiate Mycobacterium bovis from M. tuberculosis Complex (MTC) by one step amplification based on simultaneous detection of pncA 169 C > G change in M. bovis and the IS6110 present in MTC species. Our findings showed the proposed multiplex-PCR is a very useful tool for complementation in differentiating M. bovis from other cultured MTC species.
Sechi, Leonardo A.; Duprè, Ilaria; Leori, Guido; Fadda, Giovanni; Zanetti, Stefania
Amplification of a specific, 500-bp fragment from Mycobacterium bovis isolates and use of the fragment to differentiate between Mycobacterium tuberculosis and M. bovis was previously reported (J. G. Rodriguez, G. A. Meja, P. Del Portillo, M. E. Patarroyo, and L. A. Murillo, Microbiology 141:2131–2138, 1995). In the present study, 30 M. bovis isolates from Sardinian cattle were examined for the presence of this 500-bp fragment; 4 of the 30 isolates lacked the fragment. This result indicates that identification of M. bovis strains by amplification of the 500-bp sequence may lead to false-negative results. PMID:11015414
Kim, Jae Myung; Jang, Young-Boo; Jang, Yunho; Yu, So Yoon; Kim, Jiro; Moon, Oun Kyung; Jung, Suk Chan; Lee, Min Kwon; Jeong, Tae Nam
Mycobacterium (M.) bovis causes tuberculosis and has a broad host range, including humans, livestock, and wild animals. M. bovis infection of wild boar has been reported in several European countries. We report here the first case of M. bovis infection in a domesticated wild sow in Korea. Granulomatous and necrotizing lesions with small numbers of acid-fast bacilli were observed in nodules of the lung of wild sow. Furthermore, the M. bovis isolate from the wild sow had spoligotype SB0140 and a novel MIRU-VNTR allelic profile, which is not found in cattle and deer in Korea. PMID:26726026
Daly, Meighan; Diegel, Kelly L; Fitzgerald, Scott D; Schooley, Angie; Berry, Dale E; Kaneene, John B
The state of Michigan has recognized the presence of Mycobacterium bovis in its free-ranging white-tailed deer population since 1994. This endemic infection is primarily located in a 12-county area in the northeastern lower peninsula of Michigan. A statewide surveillance and eradication program of the disease has been in effect since 1994. Worldwide, Mycobacterium tuberculosis complex organisms have a known predilection toward development of antimicrobial resistance. The objective of this study was to investigate the antimicrobial susceptibility of M. bovis isolates from white-tailed deer in Michigan and detect any changes in susceptibility over time. M. bovis isolates from 2 fall hunting seasons (1999 and 2004) were used in this study. The fall season of 2004 marked the first documented case of direct transmission of M. bovis from a wild deer to a human in Michigan. Since M. bovis is a zoonotic disease, knowledge of susceptibility can expedite treatment options in humans. M. bovis isolates were obtained from 58 deer, 4 coyotes, 3 cattle, 2 raccoons, and 1 human case from the 2 years combined. Methods of susceptibility testing included 1% proportion agar plates and Bactec radiometric broth testing. M. bovis was found to be uniformly resistant to the antibiotic pyrazinamide; this resistance is common to all M. bovis isolates. No other antimicrobial resistance was found in any of the tested M. bovis isolates, which may be, in part, attributed to the lack of any significant treatment pressure in wildlife.
Ord, Rosalynn L.; Rodriguez, Marilis; Cursino-Santos, Jeny R.; Hong, Hyunryung; Singh, Manpreet; Gray, Jeremy
Apicomplexan parasites include those of the genera Plasmodium, Cryptosporidium, and Toxoplasma and those of the relatively understudied zoonotic genus Babesia. In humans, babesiosis, particularly transfusion-transmitted babesiosis, has been emerging as a major threat to public health. Like malaria, the disease pathology is a consequence of the parasitemia which develops through cyclical replication of Babesia parasites in host erythrocytes. However, there are no exoerythrocytic stages in Babesia, so targeting of the blood stage and associated proteins to directly prevent parasite invasion is the most desirable option for effective disease control. Especially promising among such molecules are the rhoptry neck proteins (RONs), whose homologs have been identified in many apicomplexan parasites. RONs are involved in the formation of the moving junction, along with AMA1, but no RON has been identified and characterized in any Babesia spp. Here we identify the RON2 proteins of Babesia divergens (BdRON2) and B. microti (BmRON2) and show that they are localized apically and that anti-BdRON2 antibodies are significant inhibitors of parasite invasion in vitro. Neither protein is immunodominant, as both proteins react only marginally with sera from infected animals. Further characterization of the direct role of both BdRON2 and BmRON2 in parasite invasion is required, but knowledge of the level of conformity of RON2 proteins within the apicomplexan phylum, particularly that of the AMA1-RON2 complex at the moving junction, along with the availability of an animal model for B. microti studies, provides a key to target this complex with a goal of preventing the erythrocytic invasion of these parasites and to further our understanding of the role of these conserved ligands in invasion. PMID:26953328
Guan, Guiquan; Ma, Miling; Liu, Aihong; Du, Pengfei; Ren, Qiaoyun; Li, Youquan; Wang, Jinming; Liu, Zhijie; Yin, Hong; Luo, Jianxun
Babesia sp. Xinjiang was isolated from a splenectomised sheep infested by Rhipicephalus sanguineus and Hylomma anatolicum anatolicum, collected from sheep and cattle in Xinjiang province. It was considered to be a novel ovine Babesia species on the basis of its morphology, pathogenicity, vector tick species and alignments of 18S ribosomal RNA (18S rRNA) and internal transcribed spacers (ITS) gene sequences. Continuous in vitro cultures of the ovine parasite were established using infected sheep blood. In RPMI 1640 medium with 7.5% sheep red blood cells (RBCs) maintained in an incubator at 37 °C and 5% CO(2), the percentage of parasitized erythrocytes (PPE) peaked at 10% in 24- and 6-well plates. It increased to 20-50% with the same culture medium but with 2.5% RBC in 75 cm(2) flasks. Two clonal lines of Babesia sp. Xinjiang were screened using the limiting dilution method. Growth characteristics of these lines in vitro were measured by a microtiter-based spectrophotometric method and from the PPE. The generation time in sheep erythrocytes was between 15.20 h and 16.27 h. Furthermore, the host range of parasite was identified with in vitro culture and in vivo infection. Erythrocytes of sheep, cattle, sika deer and humans could be invaded into by lines in vitro, but the parasites could not propagate in human erythrocytes. The parasites could not enter erythrocytes from goats in vitro. However, in vivo, only sheep could be infected by lines. Finally, a Babesia sp. Xinjiang-like parasite (which shared 99.5% identity with the original strain of Babesia sp. Xinjiang) was isolated using this in vitro culture system from 1 of 19 sheep blood samples collected from western Gansu province, China.
Barandiaran, Soledad; Martínez Vivot, Marcela; Moras, Eduardo Vicente; Cataldi, Angel Adrián; Zumárraga, Martín José
A total of 143 Mycobacterium bovis isolates of pigs, from the most productive swine area in Argentina, were typed by spoligotyping. Twenty-two different spoligotypes were identified, and 133 (93%) isolates were grouped into 12 clusters. One of them, designed SB0140, was the most frequent because it held 83 (58%) isolates. This spoligotype also grouped 362 (43%) out of 841 isolates from previously typed cattle and, thus, constitutes the most frequent in our country. In addition, 135 (94%) isolates revealed spoligotypes identical to those of cattle, showing an epidemiological link. On the other hand, there were seven novel spoligotypes, six of which were also unique since they had only one isolate each. This study aimed to identify the spoligotypes of M. bovis isolated from pigs to contribute to a better understanding of the distribution of bovine tuberculosis in the main productive area of Argentina. PMID:21547236
Zumárraga, M J; Vivot, M Martínez; Marticorena, D; Bernardelli, A; Fasán, R; Iachini, R; Cataldi, A A
In the present work, 19 Mycobacterium bovis isolates from different cats were typified by spoligotyping. We detected nine spoligotypes. There was only one cluster, which grouped 11 of the isolates (57.9%), showing the main spoligotype from cattle from Argentina. The rest of the spoligotypes presented only one isolate each. Five of them were not found in cattle, and were unique and exclusive of cats. The isolates studied show that tuberculosis of bovine origin in cats constitutes a potential public health problem in Buenos Aires region. The identification of genotypes from non-natural hosts could contribute to understand the spread of bovine tuberculosis. This is the first report showing genetic profiles of M. bovis isolates in felines from Argentina.
Ben-Chetrit, E; Wiener-Well, Y; Kashat, L; Yinnon, A M; Assous, M V
Bacteremia with Streptococcus bovis/equinus complex strains is associated with hepatobiliary disease, colorectal lesions (CL), and infective endocarditis (IE). This study addressed the clinical significance of subspecies distinction of previously designated S. bovis blood culture isolates according to the updated nomenclature. During 2002-2013, all blood culture isolates previously designated as S. bovis were recultured and identified using 16S rRNA gene sequencing and MALDI-TOF (Bruker BioTyper and Vitek MS, bioMérieux). Clinical data of patients aged ≥18 years were reviewed. A review of four recent case series was performed as well. Forty blood isolates were identified using 16S rRNA sequencing. Twenty-six bacteremic patients had S. gallolyticus ssp. pasteurianus, six had S. gallolyticus ssp. gallolyticus, two had S. gallolyticus ssp. macedonicus, and six had S. infantarius bacteremia. Species diagnosis using Vitek and bioMérieux MALDI-TOF technology was applicable in 37 and 36 samples, respectively, and was successful in all samples (100 %). Subspecies identification was confirmed in 30 (83 %) samples (as compared with 16S rRNA sequencing detection). IE was diagnosed in 22 (59 %) patients and CL in 8 (20 %) patients. Both complications were associated with all subspecies. Combining our results with those of four recent series resulted in, overall, 320 bacteremic cases, of which 88 (28 %) had CL and 66 (21 %) had IE. All 'bovis/equinus' complex subspecies were associated with CL or IE. From a clinical point of view, species diagnosis using MALDI-TOF MS should suffice to warrant consideration of transesophageal echocardiography and colonoscopy.
Background Piroplasmosis are among the most relevant diseases of domestic animals. Babesia is emerging as cause of tick-borne zoonosis worldwide and free-living animals are reservoir hosts of several zoonotic Babesia species. We investigated the epidemiology of Babesia spp. and Theileria spp. in wild ungulates and carnivores from Northern Italy to determine which of these apicomplexan species circulate in wildlife and their prevalence of infection. Methods PCR amplification of the V4 hyper-variable region of the 18S rDNA of Babesia sp./Theileria sp was carried out on spleen samples of 1036 wild animals: Roe deer Capreolus capreolus (n = 462), Red deer Cervus elaphus (n = 52), Alpine Chamois Rupicapra rupicapra (n = 36), Fallow deer Dama dama (n = 17), Wild boar Sus scrofa (n = 257), Red fox Vulpes vulpes (n = 205) and Wolf Canis lupus (n = 7). Selected positive samples were sequenced to determine the species of amplified Babesia/Theileria DNA. Results Babesia/Theileria DNA was found with a mean prevalence of 9.94% (IC95% 8.27-11.91). The only piroplasms found in carnivores was Theileria annae, which was detected in two foxes (0.98%; IC95% 0.27-3.49). Red deer showed the highest prevalence of infection (44.23%; IC95% 31.6-57.66), followed by Alpine chamois (22.22%; IC95% 11.71-38.08), Roe deer (12.55%; IC95% 9.84-15.89), and Wild boar (4.67%; IC95% 2.69-7.98). Genetic analysis identified Babesia capreoli as the most prevalent piroplasmid found in Alpine chamois, Roe deer and Red deer, followed by Babesia bigemina (found in Roe deer, Red deer and Wild boar), and the zoonotic Babesia venatorum (formerly Babesia sp. EU1) isolated from 2 Roe deer. Piroplasmids of the genus Theileria were identified in Wild boar and Red deer. Conclusions The present study offers novel insights into the role of wildlife in Babesia/Theileria epidemiology, as well as relevant information on genetic variability of piroplasmids infecting wild ungulates and
Aranaz, Alicia; de Juan, Lucía; Montero, Natalia; Sánchez, Celia; Galka, Margarita; Delso, Consuelo; Álvarez, Julio; Romero, Beatriz; Bezos, Javier; Vela, Ana I.; Briones, Victor; Mateos, Ana; Domínguez, Lucas
Mycobacterium bovis infection in wildlife and feral species is a potential source of infection for livestock and a threat to protected and endangered species. The aim of this study was to identify Spanish wild animal species infected with M. bovis through bacteriological culture and spacer oligonucleotide typing (spoligotyping) of isolates for epidemiological purposes. This study included samples from red deer (Cervus elaphus), fallow deer (Dama dama), wild boar (Sus scrofa), Iberian lynx (Lynx pardina), hare (Lepus europaeus), and cattle (Bos taurus). They were collected in several geographical areas that were selected for their unique ecological value and/or known relationships between wildlife and livestock. In the areas included in this survey, M. bovis strains with the same spoligotyping pattern were found infecting several wild species and livestock, which indicates an epidemiological link. A locally predominant spoligotype was found in these areas. Better understanding of the transmission and distribution of disease in these populations will permit more precise targeting of control measures. PMID:15184440
Vergnon-Miszczycha, Delphine; Suy, Florence; Robert, Florence; Carricajo, Anne; Fresard, Anne; Cazorla, Céline; Guglielminotti, Claire; Lucht, Frédéric; Botelho-Nevers, Elisabeth
Guillain-Barré syndrome (GBS) is an autoimmune disease that can be triggered by different infectious agents. Here we report the case of a 26-year-old Algerian woman who developed GBS associated with a Mycobacterium bovis cervical lymphadenitis. Following intravenous immunoglobulin therapy, the patient's neurologic state returned to normal after 3 months. The lymphadenitis responded more slowly to the antituberculous treatment and an excision of necrotic cervical lymph nodes had to be performed four times. Antibiotics were administered for 16 months: ethambutol was stopped after 2 months, and rifampicin and isoniazid pursued for 14 months. An extensive etiological investigation showed that, in this case, the only likely infectious trigger GBS was the concomitant M. bovis infection. To our knowledge, this is the first report of GBS triggered by M. bovis. We performed a literature review revealing that the association between tuberculosis and Guillain-Barré syndrome is very rare (only seven cases previously reported) but is not coincidental. Physicians should be aware that tuberculosis can be a cause of GBS.
Hermosilla, Carlos; Ruiz, Antonio; Taubert, Anja
Apicomplexan parasites are obligate intracellular protozoans and are well recognized modulators of the host cell machinery on varying levels such as host cell metabolism, MHC expression, cell cycle, or apoptosis in order to guarantee their intracellular development and survival. One of the most thoroughly examined apicomplexan pathogens demonstrating a potent manipulative capacity with respect to various host cell functions is Toxoplasma gondii, a protozoon exhibiting rapid intracellular development with small meronts in any nucleated cell, almost irrespective of the cell type or host origin. In contrast, Eimeria bovis merogony I is host- and cell type-restricted and occurs exclusively in bovine endothelial host cells. Furthermore, as a peculiarity, intracellular E. bovis meront I development is a long-lasting process (up to 3 weeks), leading to the formation of huge macromeronts of up to 300 μm in size, containing up to 120,000 merozoites I as offspring. In consequence, the necessity for intense host cell modulation to support this particular development appears even more pressing than in other apicomplexan parasite cases. Here we review the data currently available on E. bovis-host cell interactions, indicating the intriguing capacity of this protozoan to exploit and utilize its host cell for its own benefit.
Herrera-Sánchez, Diana Andrea; Castilla-Rodríguez, Jaisel Luz; Castrejón-Vázquez, María Isabel; Vargas-Camaño, María Eugenia; Medina-Torres, Edgar Alejandro; Blancas-Galicia, Lizbeth; Espinosa-Padilla, Sara Elva
Common variable immunodeficiency (CVID) is an heterogeneous group of disorders characterized by impaired antibody production. It shows a wide spectrum of manifestations including severe and recurrent respiratory infections (Streptococcus pneumoniae, Haemophilus) and gastrointestinal (Campylobacter jejuni, rotavirus and Giardia lamblia). Viral infections caused by herpes zoster, cytomegalovirus (CMV) and hepatitis C are rare. The opportunistic agents such as CMV, Pneumocystis jirovecii, cryptococcus and atypical mycobacteria have been reported as isolated cases. This paper reports the case of a 38-year-old female patient, who began six years before with weight loss of 7 kg in six months, fatigue, weakness, sweating, fever and abdominal pain. Furthermore, patient had intestinal obstruction and abdominal CT showed mesenteric lymph growth. The mesenteric lymph node biopsy revealed positives Mycobacterium PCR, Ziehl-Neelsen staining and culture for M. bovis. In the laparotomy postoperative period was complicated with nosocomial pneumonia, requiring mechanical ventilation and tracheostomy. Two years later, she developed right renal abscess that required surgical drainage, once again with a positive culture for Mycobacterium bovis. She was referred to highly specialized hospital and we documented panhypogammaglobulinemia and lymphopenia. Secondary causes of hypogammaglobulinemia were ruled out and common variable immunodeficiency (CVID) was confirmed, we started IVIG replacement. Four years later she developed mixed cellularity Hodgkin's lymphoma. Until today she continues with IVIG and chemotherapy. This report of a patient with CVID and Mycobacterium bovis infection, a unusual association, shows the cellular immunity susceptibility in this immunodeficiency, additional to the humoral defect.
Rocha, Vivianne Cambuí Figueiredo; Figueiredo, Salomão Cambuí de; Rosales, Cesar Alejandro Rodriguez; Porto, Camila Dias; Sequeira, Julio Lopes; Neto, José Soares Ferreira; Paes, Antônio Carlos; Salgado, Vanessa Riesz
Tuberculosis (TB) is a chronic disease caused by bacteria belonging to the Mycobacterium tuberculosis complex (MtbC). This disease rarely affects dogs. Canine infections are usually caused by M. tuberculosis. Mycobacterium bovis infections are rare in dogs and associated with consumption of raw milk or contaminated products. Here, we report a Boxer dog who had a M. bovis infection and was admitted to a Brazilian veterinary hospital with a presumptive diagnosis of chronic ehrlichiosis. Despite receiving treatment for chronic ehrlichiosis, it progressed to death. TB was diagnosed during post-mortem examinations using histopathological analysis. Ziehl-Neelsen staining revealed acid-fast bacilli in the kidneys, liver, mesentery, and a mass adhered to the liver. Further, PCR-restriction analysis was performed to identify mycobacteria in the samples. A restriction profile compatible with MtbC was found in the lungs. In addition, PCR-based MtbC typing deletions at different loci of chromosome 9 enabled the identification of M. bovis in the lungs. Therefore, it is very essential to perform differential diagnosis of TB in dogs with non-specific clinical signs and who do not respond to treatment, particularly those who had been in contact with TB-infected cattle or owners. Further, we highlight the use of molecular methods for the identification of bacilli, improving the diagnosis and aiding epidemiological studies.
Van Heerden, J
Babesia canis was successfully transmitted from the domestic dog to 3 wild dogs Lycaon pictus and 4 black-backed jackals Canis mesomelas. Both wild dogs and black-backed jackals showed no clinical signs or clinical pathological evidence of disease. Trophozoites of Babesia canis were found in peripheral blood smears from all experimental animals. The disease was also successfully transmitted from both black-backed jackals and wild dogs to the domestic dog.
Rar, V A; Epikhina, T I; Suntsova, O V; Kozlova, I V; Lisak, O V; Pukhovskaya, N M; Vysochina, N P; Ivanov, L I; Tikunova, N V
To study Babesia diversity in Ixodid ticks in Russia, Ixodes persulcatus, Haemaphysalis japonica, Haemaphysalisconcinna, Dermacentor silvarum, and Dermacentor nuttalli ticks collected in the Far East and Baikal region were assayed for the presence of Babesia spp. using nested PCR. In total, Babesia DNA was detected in 30 of the 1125 (2.7%) I. persulcatus, 17 of the 573 (3.0%) H. concinna, and 12 of the 543 (2.2%) H. japonica but was undetectable in any of the 294 analyzed Dermacentor spp. Partial 18S rRNA gene sequences were determined for all of the positive samples. Among the positive ticks, nine I. persulcatus were infected by Babesia microti 'US'-type, five I. persulcatus were infected by Babesia divergens-like parasites, and 11 I. persulcatus were infected by Babesia venatorum. For all three of these species, the determined 18S rRNA gene sequences were identical to those of the Babesia genetic variants found previously in I. persulcatus in Russia. In addition, five I. persulcatus from the Baikal region and all of the positive Haemaphysalis spp. ticks carried 13 different sequence variants of Babesia sensu stricto belonging to distinct phylogenetic clusters. Babesia spp. from 29 ticks of different species collected in distinct locations belonged to the cluster of cattle and ovine parasites (Babesia crassa, Babesiamajor, Babesiamotasi, Babesiabigemina, etc.). Babesia spp. from four H. japonica ticks in the Far East belonged to the cluster formed by parasites of carnivores. One more Babesia sequence variant detected in an I. persulcatus tick from the Baikal region belonged to the cluster formed by parasites of cattle and wild cervids (B. divergens, Babesiacapreoli, B. venatorum, Babesiaodocoilei, etc.).
Clarridge, J E; Attorri, S M; Zhang, Q; Bartell, J
We characterized 22 human clinical strains of Streptococcus bovis by genotypic (16S rRNA gene sequence analysis [MicroSeq]; Applied Biosystems, Foster City, Calif.) and phenotypic (API 20 Strep and Rapid ID32 Strep systems (bioMerieux Vitek, Hazelton, Mo.) methods. The strains, isolated from blood, cerebrospinal fluid (CSF), and urine, formed two distinct 16S ribosomal DNA sequence clusters. Three strains which were associated with endocarditis urinary tract infection (UTI), and sepsis clustered with the S. bovis type strain ATCC 33317 (cluster 1); other closely related type strains were S. equinus and S. infantarius. Nineteen strains clustered at a distance of about 2.5% dissimilarity to the S. bovis type strain (cluster 2) and were associated with central nervous system (CNS) disease in addition to endocarditis, UTI, and sepsis. All strains were distinct from S. gallolyticus. Within cluster 2, a single strain grouped with ATCC strain 43143 (cluster 2a) and may be phenotypically distinct. All the other strains formed a second subgroup (cluster 2b) that was biochemically similar to S. bovis biotype II/2 (mannitol negative and beta galactosidase, alpha galactosidase, beta glucuronidase, and trehalose positive). The API 20 Strep system identified isolates of cluster 2b as S. bovis biotype II/2, those of cluster 1 as S. bovis biotype II/1, and that of cluster 2a as S. bovis biotype I. There was an excellent correlation of biotype and genotype: S. bovis biotype II/2 isolates form a separate genospecies distinct from the S. bovis, S. gallolyticus, and S. infantarius type strains and are the most common isolates in adult males.
Mycobacterium bovis is the cause of tuberculosis in most animal species, including cattle and is a serious zoonotic pathogen. In humans, M. bovis infection can result in disease clinically indistinguishable from that caused by Mycobacterium tuberculosis, the cause of most tuberculosis in humans. Reg...
Branger, Maxime; Hauer, Amandine; Michelet, Lorraine; Karoui, Claudine; Cochard, Thierry; De Cruz, Krystel; Henault, Sylvie
Mycobacterium bovis is the etiologic agent of bovine tuberculosis, a chronic infectious disease, affecting livestock, wild animals, and sometimes humans. We report the draft genome sequence of a Mycobacterium bovis strain isolated from wild boar of spoligotype SB0120 (or BCG-like) also present in wildlife-livestock multi-host systems. PMID:27834714
Mycoplasma bovis is a primary agent of mastitis, pneumonia and arthritis in cattle and is the bacterium isolated most frequently from the polymicrobial syndrome known as bovine respiratory disease complex (BRDC). Recently, M. bovis has emerged as a significant problem in bison, causing necrotic pha...
Mycoplasma bovis is a primary agent of mastitis, pneumonia and arthritis in cattle and is the bacterium isolated most frequently from the polymicrobial syndrome known as bovine respiratory disease complex (BRDC). Recently, M. bovis has emerged as a significant health problem in bison, causing necro...
Wildlife reservoirs of Mycobacterium bovis represent serious obstacles to the eradication of tuberculosis in domestic livestock. In Michigan, USA tuberculous white-tailed deer transmit M. bovis to cattle. One approach in dealing with this wildlife reservoir is to vaccinate deer in order to interrupt...
Manuel, Christopher; Pugazhenthi, Umarani; Spiegel, Shannon; Leszczynski, Jori
Rodent health-monitoring programs based on sampling an IVC system's exhaust air dust (EAD) has enhanced and evenreplaced traditional sentinels for some rodent pathogens. EAD testing by qPCR assay is an optimal surveillance methodfor the rapid detection of Corynebacterium bovis-infected immunodeficient mice. Here we demonstrate that an active EADsurveillance program for C. bovis can be used to maintain nude mice C. bovis-free after the transition from historically enzootically infected colonies. During 3 events over 3 y, rapid detection of infection, elimination of infected mice, aggressivequarantine measures, and local decontamination prevented the spread of C. bovis within 2 barrier rooms. In total, 4 cages ofinfected nude mice were identified and removed, preventing the spread of infection to 469 other cages of immunodeficientmice. In addition, we present data regarding a refinement to EAD testing which enables row-specific surveillance of an IVCrack. This technique systemically decreases the amount of testing required to locate an individually infected cage. Due to ourability to rapidly detect and localize an infected cage, we were able to investigate the route of C. bovis introduction into ourbarrier rooms. Our epidemiologic investigation suggested that the transmission of C. bovis occurred through contaminated,cryopreserved, patient-derived xenograft tumor tissue. This previously unknown source of C. bovis can infect mice used topropagate these tumors. Together, these data demonstrate that a remediation program that combines rapid detection, testand-cull, and local decontamination under quarantine conditions can eliminate C. bovis from a mouse colony.
Recently, several serologic tests designed to detect immunodominant antibodies to M. bovis antigens (e.g., MPB83, MPB70, ESAT-6, and CFP10) have emerged for potential use with samples from cattle. Of these, a commercial ELISA to MPB83/MPB70 (M. bovis antibody ELISA) has gained approval for use in ca...
Díez-Delgado, Iratxe; Contreras, Marinela; Vicente, Joaquín; Cabezas-Cruz, Alejandro; Manrique, Marina; Tobes, Raquel; López, Vladimir; Romero, Beatriz; Domínguez, Lucas; Garrido, Joseba M.; Gortazar, Christian
Here we report the complete genome sequences of field isolates of Mycobacterium bovis and the related mycobacterial species, Mycobacterium caprae. The genomes of three M. bovis (MB1, MB3, MB4) and one M. caprae (MB2) field isolates with different virulence, prevalence, and host distribution phenotypes were sequenced. PMID:26112781
Mycobacterium tuberculosis and Mycobacterium bovis are responsible for tuberculosis in humans or animals, respectively. Both species are closely related and belong to the Mycobacterium tuberculosis complex (MTC). M. tuberculosis is the most ancient species from which M. bovis and the other members o...
Szewzyk, R; Svenson, S B; Hoffner, S E; Bölske, G; Wahlström, H; Englund, L; Engvall, A; Källenius, G
Forty-nine isolates of Mycobacterium bovis from humans and animals in Sweden were analyzed by restriction fragment length polymorphism (RFLP) patterns probed by the insertion element IS6110. Most isolates had patterns indicating the presence of only one or two genomic copies of the IS6110 insertion element. This simple type of pattern was found in all human isolates. In contrast, isolates from M. bovis infections in five herds of farmed deer in Sweden showed a specific RFLP pattern with seven bands, indicating seven copies of the IS6110 sequence. In 1958, Sweden was declared free from M. bovis in cattle. However, in 1987, M. bovis was reintroduced with imported farmed deer, and since 1991, 11 outbreaks in deer herds, but not in other livestock or wildlife, have been diagnosed. Continued RFLP studies of the new Swedish M. bovis isolates can reveal possible transmission of this deer strain to other animals or humans. PMID:8586698
Arcangioli, Marie-Anne; Duet, Arnaud; Meyer, Gilles; Dernburg, Ann; Bézille, Pierre; Poumarat, François; Le Grand, Dominique
To assess the prevalence and relative importance of Mycoplasma bovis among the pathological agents implicated in bovine respiratory disease (BRD), we surveyed 135 veal calves from nine feedlots in eastern France during naturally occurring outbreaks of respiratory disease. Occurrence of respiratory pathogens, M. bovis, bovine viral diarrhoea (BVD) virus, bovine respiratory syncytial (BRS) virus and parainfluenza-3 (PI3) virus was investigated by seroconversion and isolation of bacteria and viruses from broncho-alveolar fluids. M. bovis and pathogenic respiratory bacteria were isolated in eight of the nine feedlots, and from 106 and 32, respectively, of the 135 tested animals. Seroconversion to PI3 virus occurred in four lots. BVD and BRS viruses were detected in eight and one lot, respectively. M. bovis was the most frequently isolated aetiologic agent in these BRD outbreaks, spreading early and widely throughout the affected units (60-100% rate of isolation and seroconversion). These results stress the importance of M. bovis in the BRD complex.
Zumárraga, Martín J; Meikle, Virginia; Bernardelli, Amelia; Abdala, Alejandro; Tarabla, Hector; Romano, María I; Cataldi, Angel
The confirmatory diagnosis of Mycobacterium bovis (M. bovis) in animal samples is carried out by culture in Stonebrink media. However, culture is very slow because of the extremely long duplication time of the bacillus and difficult because of the scarcity of bacilli in diagnostic samples. This study describes the development of a single-tube touch-down polymerase chain reaction (PCR) protocol for the detection of M. bovis using primers that target the IS6110 element. Spiked water and milk as well as routine diagnostic samples (milk and nasal swabs) from M. bovis-positive cattle were tested. This protocol allows the rapid and sensitive detection of M. bovis in bovine samples by enhancing the sensitivity of standard PCR amplification.
Brooks, B W; Barnum, D A; Meek, A H
An observational study of Corynebacterium bovis was conducted in 74 Ontario dairy herds. The levels of infection with C. bovis were 19.9, 36.2 and 85.6% at the quarter, cow and herd level, respectively. Teat disinfection was found to be the variable best able to distinguish between herds with a high or low C. bovis quarter infection rate. Mean total milk somatic cell counts for 1103 quarters and 107 cows infected with only C. bovis ranged between 150,000 and 200,000/mL and were significantly higher than for uninfected quarters or cows. The rate of infection with mastitis pathogens was not significantly different in quarters previously colonized with only C. bovis compared to previously uninfected quarters. PMID:6831308
Cézanne, Rita; Mrowietz, Naike; Eigner, Barbara; Duscher, Georg Gerhard; Glawischnig, Walter; Fuehrer, Hans-Peter
Wild ungulates may act as reservoirs of various vector borne pathogens that can infect humans and domestic animals. In the present study, blood samples from 196 red deer (Cervus elaphus) from Western Austria (Vorarlberg, Tyrol and Salzburg) were collected on filter paper and tested for Anaplasmataceae, Piroplasmida, Rickettsia and filarioid helminths using molecular tools. Babesia divergens was detected in ten (5.1%) and Anaplasma phagocytophilum in three (1.5%) of the 196 samples. Filarioid helminths, Rickettsia spp. and Theileria spp. were not detected. These findings indicate that red deer may serve as reservoirs of Babesia divergens and Anaplasma phagocytophilum in Western Austria. Further investigations are needed to assess the presence of these pathogens in ticks in this geographical region, and the significance of these pathogens in both animals and humans.
Yabsley, Michael J.; Work, Thierry M.; Rameyer, Robert A.
The phylogenetic relationship of avian Babesia with other piroplasms remains unclear, mainly because of a lack of objective criteria such as molecular phylogenetics. In this study, our objective was to sequence the entire 18S, ITS-1, 5.8S, and ITS-2 regions of the rRNA gene and partial ß-tubulin gene of B. poelea, first described from brown boobies (Sula leucogaster) from the central Pacific, and compare them to those of other piroplasms. Phylogenetic analyses of the entire 18S rRNA gene sequence revealed that B. poelea belonged to the clade of piroplasms previously detected in humans, domestic dogs, and wild ungulates in the western United States. The entire ITS-1, 5.8S, ITS-2, and partial ß-tubulin gene sequence shared conserved regions with previously described Babesia and Theileria species. The intron of the ß-tubulin gene was 45 bp. This is the first molecular characterization of an avian piroplasm.
Yabsley, M.J.; Work, T.M.; Rameyer, R.A.
The phylogenetic relationship of avian Babesia with other piroplasms remains unclear, mainly because of a lack of objective criteria such as molecular phylogenetics. In this study, our objective was to sequence the entire 18S, ITS-1, 5.8S, and ITS-2 regions of the rRNA gene and partial ??-tubulin gene of B. poelea, first described from brown boobies (Sula leucogaster) from the central Pacific, and compare them to those of other piroplasms. Phylogenetic analyses of the entire 18S rRNA gene sequence revealed that B. poelea belonged to the clade of piroplasms previously detected in humans, domestic dogs, and wild ungulates in the western United States. The entire ITS-1, 5.8S, ITS-2, and partial ??-tubulin gene sequence shared conserved regions with previously described Babesia and Theileria species. The intron of the ??-tubulin gene was 45 bp. This is the first molecular characterization of an avian piroplasm. ?? American Society of Parasitologists 2006.
Usatii, Natalia; Khachatrian, Aelita; Stratidis, John
This article describes the case of spontaneous splenic rupture as a rare complication of infection with Babesia species. We will discuss the symptomatology that this disease could present along with both surgical and non-surgical management approaches. Babesia infection often presents with mild to moderate symptoms, but can rapidly progress to significant injury including splenic rupture. The first case reported in a medical journal was in 2007. Treatment usually involves a two-drug regimen; clindamycin plus quinine, or atovaquone plus azithromycin (as in our patient). If hemodynamic stability is present, a primary non-surgical treatment may be especially beneficial since splenectomy may worsen optimal immunologic function and the infection itself. PMID:26839774
Bednarska, Malgorzata; Bajer, Anna; Drozdowska, Anna; Mierzejewska, Ewa J.; Tolkacz, Katarzyna; Welc-Falęciak, Renata
Babesia spp. (Apicomplexa, Piroplasmida) are obligate parasites of many species of mammals, causing a malaria-like infection- babesiosis. Three routes of Babesia infection have been recognized to date. The main route is by a tick bite, the second is via blood transfusion. The third, vertical route of infection is poorly recognized and understood. Our study focused on vertical transmission of B. microti in a well-established mouse model. We assessed the success of this route of infection in BALB/c mice with acute and chronic infections of B. microti. In experimental groups, females were mated on the 1st day of Babesia infection (Group G0); on the 28th day post infection (dpi) in the post- acute phase of the parasite infection (G28); and on the 90th and 150th dpi (G90 and G150 group, respectively), in the chronic phase of the parasite infection. Pups were obtained from 58% of females mated in the post-acute phase (G28) and from 33% of females in groups G90 and G150. Mice mated in the pre-acute phase of infection (G0) did not deliver pups. Congenital B. microti infections were detected by PCR amplification of Babesia 18S rDNA in almost all pups (96%) from the experimental groups G28, G90 and G150. Parasitaemia in the F1 generation was low and varied between 0.01–0.001%. Vertical transmission of B. microti was demonstrated for the first time in BALB/c mice. PMID:26372043
Taubert, Anja; Behrendt, Jan Hillern; Sühwold, Anke; Zahner, Horst; Hermosilla, Carlos
Innate immune reactions conducted by macrophages may affect the outcome of primary infections and are crucial for the transition to adaptive immune responses. In bovine coccidiosis little is known on early monocyte/macrophage-mediated responses. We therefore investigated in vivo, in vitro and ex vivo reactions of monocytes and macrophages against Eimeria bovis, one of the most pathogenic Eimeria species in cattle. Macrophages significantly infiltrate the gut mucosa of E. bovis-infected calves, particularly after challenge infection. Furthermore, peripheral monocytes of infected animals, as precursor cells of macrophages, exhibited enhanced ex vivo phagocytic and oxidative burst activities. Enhanced levels of both activities were found early after infection and towards the end of first merogony. In vitro exposure of macrophages to sporozoites led to phagocytosis of the pathogen, whilst monocytes failed to do so. Phagocytosis occurred independently of the viability of the sporozoites, indicating that active invasion by the parasites was negligible. Phagocytosis occurred in the absence of immune serum, but could clearly be enhanced by addition of immune serum, suggesting macrophage-derived antibody-dependent cytotoxicity. Furthermore, co-culture of macrophages with sporozoites and stimulation with merozoite I antigen induced distinct levels of cytokine and chemokine gene transcription. Thus, the transcription of genes encoding for IFN-gamma, IL-12, TNF-alpha, IL-6, CXCL1, CXCL8, CXCL10 and COX-2 was upregulated after sporozoite encounter. In contrast, soluble merozoite I antigen only induced the gene transcription of IL-6 and IL-12 and failed to upregulate IFN-gamma and TNF-alpha gene transcripts. In monocytes, IFN-gamma and CXCL10 were found upregulated, all other immunoregulatory molecules tested were not affected. In summary, our results strongly suggest that macrophage-mediated, innate immune reactions play an important role in the early immune response to E
Lang, Mirjam; Kann, Michael; Zahner, Horst; Taubert, Anja; Hermosilla, Carlos
Sophisticated evasion strategies of obligate intracellular parasites, in particular prevention of host cell apoptosis, are necessary to ensure successful replication. To study the ability of Eimeria bovis in this regard, in vitro experiments were performed applying bovine foetal gastrointestinal cells (BFGC), bovine umbilical vein endothelial cells (BUVEC) and African green monkey kidney cells (VERO) as host cells. BUVEC and BFGC allow maturation of sporozoites to macromeronts, in VERO cells sporozoites survive for weeks without showing further development. In highly infected BUVEC monolayers, infected cells survived until merozoite release whereas uninfected cells underwent apoptosis. Light microscopy and TUNEL assays performed 3-10 days p.i. showed that, within infected BFGC and VERO cell monolayers, uninfected cells underwent programmed cell death after application of various inducers of apoptosis, whereas infected cells survived. Incidentally, the anti-apoptotic efficacies in infected cells were independent of the drugs and the host cell type. We could not demonstrate significant differences between infected and uninfected cells after colchicin treatment in terms of translation of phosphatidylserines to the host cell surface, caspase 3 activity and cytochrome c release, probably since obtainable infection rates were too low. However, we could show by laser scanning confocal microscopy on single cell levels that the expression of the anti-apoptotic factors cellular Flice inhibitory protein (c-FLIP) and cellular inhibition of apoptosis protein 1 (c-IAP1) were enhanced in E. bovis infected cells after application of colchicin, in the latter case also in non-infected cells directly neighbouring infected ones. Our data show that E. bovis protects its host cell from apoptosis by increasing expression of c-IAP1 and c-FLIP.
Tsuji, Naotoshi; Miyoshi, Takeharu; Battsetseg, Badger; Matsuo, Tomohide; Xuan, Xuenan; Fujisaki, Kozo
Vector ticks possess a unique system that enables them to digest large amounts of host blood and to transmit various animal and human pathogens, suggesting the existence of evolutionally acquired proteolytic mechanisms. We report here the molecular and reverse genetic characterization of a multifunctional cysteine protease, longipain, from the babesial parasite vector tick Haemaphysalis longicornis. Longipain shares structural similarity with papain-family cysteine proteases obtained from invertebrates and vertebrates. Endogenous longipain was mainly expressed in the midgut epithelium and was specifically localized at lysosomal vacuoles and possibly released into the lumen. Its expression was up-regulated by host blood feeding. Enzymatic functional assays using in vitro and in vivo substrates revealed that longipain hydrolysis occurs over a broad range of pH and temperature. Haemoparasiticidal assays showed that longipain dose-dependently killed tick-borne Babesia parasites, and its babesiacidal effect occurred via specific adherence to the parasite membranes. Disruption of endogenous longipain by RNA interference revealed that longipain is involved in the digestion of the host blood meal. In addition, the knockdown ticks contained an increased number of parasites, suggesting that longipain exerts a killing effect against the midgut-stage Babesia parasites in ticks. Our results suggest that longipain is essential for tick survival, and may have a role in controlling the transmission of tick-transmittable Babesia parasites. PMID:18483546
Simking, Patcharathorn; Wongnakphet, Sirichai; Stich, Roger W; Jittapalapong, Sathaporn
The combination of a rapidly growing stray animal population and the lack of animal control in Bangkok has resulted in a unique opportunity to evaluate the potential role of companion animals as sentinels and reservoirs of infectious diseases, including several of those caused by vector-borne parasites. The purpose of this study was to determine the prevalence and factors associated with the distribution of Babesia species infections among stray cats in Bangkok. Blood samples were collected from 1490 stray cats residing in 140 monasteries of 50 metropolitan districts of Bangkok, and assayed with light microscopy and PCR for evidence of Babesia spp. Pear-shaped merozoites were observed microscopically from two (0.13%) of these cats, while a nested 18S rDNA-based PCR assay detected babesial infections in 21 (1.4%) of the cats tested. The prevalence of infection was significantly different between sexes (p<0.05), and PCR-positive cats were found in 30% (15/50) of the districts surveyed. All 21 amplicon sequences were identical, and were determined to be closest to that reported for B. vogeli (98% identity). These results represent the first molecular confirmation that a Babesia sp. is enzootic among stray cat populations in Thailand, and suggest that the presence of pet companion animals could be a risk factor for exposure of stray cats to vector-borne parasites.
Cardoso, L; Cortes, H C E; Reis, A; Rodrigues, P; Simões, M; Lopes, A P; Vila-Viçosa, M J; Talmi-Frank, D; Eyal, O; Solano-Gallego, L; Baneth, G
The prevalence of piroplasm (order Piroplasmida) infection was assessed in blood and bone marrow samples from 91 red foxes (Vulpes vulpes) from northern, central and southern Portugal by means of molecular methods. PCR for the 18S rRNA gene of Babesia spp. followed by sequencing revealed 63 foxes positive for the Babesia microti-like piroplasm (syn. Theileria annae) (69.2%; 95% confidence interval [CI]: 58.7-78.5%) and one fox positive for Babesia canis (1.1%; 95% CI: 0.0-6.0%). Positivity to the B. microti-like piroplasm or B. canis in 43 blood samples (83.7%) was significantly higher (p<0.001) than in 43 paired bone marrow samples (20.9%). There were no statistically significant differences in the prevalence of infection between genders (p=0.219) or age groups (<2 years vs. ≥ 2 years) (p=1.0). This is the first report of the B. microti-like piroplasm in foxes from Portugal as well as the first report on detection by PCR and genotyping of B. canis in a red fox worldwide. A natural cycle of the B. microti-like piroplasm is suggested in red fox populations based on the high prevalence of the protozoan. Red foxes might be a reservoir of the B. microti-like piroplasm and a source of infection to dogs.
Rodriguez, Marilis; Alhassan, Andy; Ord, Rosalynn L.; Cursino-Santos, Jeny R.; Singh, Manpreet; Gray, Jeremy; Lobo, Cheryl A.
Human babesiosis is caused by one of several babesial species transmitted by ixodid ticks that have distinct geographical distributions based on the presence of competent animal hosts. The pathology of babesiosis, like malaria, is a consequence of the parasitaemia which develops through the cyclical replication of Babesia parasites in a patient's red blood cells, though symptoms typically are nonspecific. We have identified the gene encoding Rhoptry-Associated Protein −1 (RAP-1) from a human isolate of B. divergens, Rouen1987 and characterized its protein product at the molecular and cellular level. Consistent with other Babesia RAP-1 homologues, BdRAP-1 is expressed as a 46 kDa protein in the parasite rhoptries, suggesting a possible role in red cell invasion. Native BdRAP-1 binds to an unidentified red cell receptor(s) that appears to be non-sialylated and non-proteinacious in nature, but we do not find significant reduction in growth with anti-rRAP1 antibodies in vitro, highlighting the possibility the B. divergens is able to use alternative pathways for invasion, or there is an alternative, complementary, role for BdRAP-1 during the invasion process. As it is the parasite's ability to recognize and then invade host cells which is central to clinical disease, characterising and understanding the role of Babesia-derived proteins involved in these steps are of great interest for the development of an effective prophylaxis. PMID:25226276
Cosivi, O.; Grange, J. M.; Daborn, C. J.; Raviglione, M. C.; Fujikura, T.; Cousins, D.; Robinson, R. A.; Huchzermeyer, H. F.; de Kantor, I.; Meslin, F. X.
The World Health Organization (WHO) estimates that human tuberculosis (TB) incidence and deaths for 1990 to 1999 will be 88 million and 30 million, respectively, with most cases in developing countries. Zoonotic TB (caused by Mycobacterium bovis) is present in animals in most developing countries where surveillance and control activities are often inadequate or unavailable; therefore, many epidemiologic and public health aspects of infection remain largely unknown. We review available information on zoonotic TB in developing countries, analyze risk factors that may play a role in the disease, review recent WHO activities, and recommend actions to assess the magnitude of the problem and control the disease in humans and animals. PMID:9452399
Poumarat, F; Martel, J L
The in vitro activity of 15 antibiotics was tested with 30-90 Mycoplasma bovis representative strains of bovine lung pathology in France. The distribution of minimal inhibitory concentration (MIC) is homogeneous with low values for spectinomycin, lincomycin, tylosin, gentamicin and baytril, intermediate for chloramphenicol and neomycin, high for nalidixic acid, Flumequine and erythromycin. The MIC distribution is heterogeneous with intermediate values for spiramycin and tetracyclines, and high values for streptomycin. For the later antibiotics, the heterogeneity of the susceptibility suggests a mechanism of acquired resistance.
Singla, Lachhman Das; Sumbria, Deepak; Mandhotra, Ajay; Bal, M S; Kaur, Paramjit
There are few published studies on various vector borne diseases of dogs in India and most depict clinical infection in dogs, diagnosed by observation of the haemopathogens in stained blood smears. This study provides the first report regarding molecular confirmation and ancestral relationship analysis of blood smears positive cases of assorted haemopathogens in Punjab province of India. On blood smear examination, haemopathogens were observed in 124 out of 778 (15.95%, 95% CI: 13.53- 18.68) blood smears. Further polymerase chain reactions (PCR) was used on bloods smear positive cases to validate the results. Out of 778 blood samples, Babesia gibsoni was most common parasite infecting dogs (15.04%, 95% CI: 12.7-17.72), followed by Ehrlichia canis (0.39%, 95% CI: 0.0-1.13), infection of Babesia vogeli and Hepatozoon canis was same (0.26%, 95% CI: 0.0-0.9). Among various risk factors studied (age, sex, season), prevalence of infection was non-significantly higher in 1-2 year of age group (19.88%, 95% CI: 14.45-26.71), regarding sex same prevalence was recorded (15.94%), and chances of infection was highest in pre-monsoon i.e. summer (18.26%, 95% CI: 14.49-22.76). Phylogenetic analysis revealed ancestral background of Ludhiana isolates of B. vogeli, B. gibsoni, H. canis, and E. canis with the isolates of Philippines, Mongolia and Tunisia.
Gottlieb, Juliana; André, Marcos Rogério; Soares, João Fábio; Gonçalves, Luiz Ricardo; Tonial de Oliveira, Mateus; Costa, Marcio Machado; Labruna, Marcelo Bahia; Bortolini, Carlos Eduardo; Machado, Rosangela Zacarias; Vieira, Maria Isabel Botelho
Pathogens transmitted by ticks are an emerging problem worldwide, this study aimed to diagnose the causal agents of infection in dogs presenting suspected hemoparasitoses. Fifty-eight dogs with clinical signs such as depression, hemorrhagic diathesis and fever were evaluated regarding clinical presentation, hemogram, blood smears and serological tests, using the indirect immunofluorescence method for the agents Babesia vogeli and Ehrlichia canis and conventional PCR for Babesia spp. (gene 18S rRNA), Rangelia vitalii (gene 18S rRNA) and Ehrlichia spp. (gene dsb). Five (8.6%) of the 58 dogs were serologically positive for Babesia spp. and three (5.1%) for E. canis. Four dogs (6.8%) were positive for R. vitalii through the molecular diagnosis. The PCR products were sequenced and the DNA from R. vitalii was found to be 99% genetically identical to samples of R. vitalii that had been isolated in Brazil. No presence of Babesia spp. or E. canis was observed through PCR on the dogs evaluated here. The results indicate the presence of R. vitalii and exposure to Babesia spp. and Ehrlichia spp. among the dogs analyzed.
Manuel, Christopher; Bagby, Stacey; Reisinger, Julie; Pugazhenthi, Umarani; Pitts, Todd; Keysar, Stephen; Arcaroli, John; Leszczynski, Jori
Human patient-derived xenograft (PDX) tumors, propagated in immunodeficient mice, are rapidly growing in use as amodelfor cancer research. Horizontal transfer between mice, without in vitro cell culture, allows these tumors to retainmany of their unique characteristics from their individual patient of origin. However, the immunodeficient mouse strainsused to grow these tumors are susceptible to numerous opportunistic pathogens, including Corynebacterium bovis. At ourinstitution, 2 in vivo tumor banks of PDX tumors had been maintained within nude mouse colonies enzootically infectedwith C. bovis. Elimination of C. bovis from these colonies required the aseptic harvest and horizontal transfer of tumor tissue between infected and naïve recipient mice without cross-contamination. Out of necessity, we developed a standard operating procedure using enhancements to traditional aseptic surgical technique with concurrent application of both procedural and physical barriers to prevent C. bovis transmission. By using these methods, all 61 unique PDX tumor models were successfullyharvested from C. bovis-infected mice and transferred into recipient mice without transmission of infection. Our datademonstrate that, in situations where C. bovis-free colonies can be established and maintained, this procedure can successfullybe used to eliminate C. bovis from an in vivo tumor bank of valuable PDX tumors.
Manuel, Christopher A; Bagby, Stacey M; Reisinger, Julie A; Pugazhenthi, Umarani; Pitts, Todd M; Keysar, Stephen B; Arcaroli, John J; Leszczynski, Jori K
Human patient-derived xenograft (PDX) tumors, propagated in immunodeficient mice, are rapidly growing in use as a model for cancer research. Horizontal transfer between mice, without in vitro cell culture, allows these tumors to retain many of their unique characteristics from their individual patient of origin. However, the immunodeficient mouse strains used to grow these tumors are susceptible to numerous opportunistic pathogens, including Corynebacterium bovis. At our institution, 2 in vivo tumor banks of PDX tumors had been maintained within nude mouse colonies enzootically infected with C. bovis. Elimination of C. bovis from these colonies required the aseptic harvest and horizontal transfer of tumor tissue between infected and naïve recipient mice without cross-contamination. Out of necessity, we developed a standard operating procedure using enhancements to traditional aseptic surgical technique with concurrent application of both procedural and physical barriers to prevent C. bovis transmission. By using these methods, all 61 unique PDX tumor models were successfully harvested from C. bovis-infected mice and transferred into recipient mice without transmission of infection. Our data demonstrate that, in situations where C. bovis-free colonies can be established and maintained, this procedure can successfully be used to eliminate C. bovis from an in vivo tumor bank of valuable PDX tumors.
Woodroffe, Rosie; Donnelly, Christl A; Wei, Gao; Cox, D R; Bourne, F John; Burke, Terry; Butlin, Roger K; Cheeseman, C L; Gettinby, George; Gilks, Peter; Hedges, Simon; Jenkins, Helen E; Johnston, W Thomas; McInerney, John P; Morrison, W Ivan; Pope, Lisa C
1. In most social animals, the prevalence of directly transmitted pathogens increases in larger groups and at higher population densities. Such patterns are predicted by models of Mycobacterium bovis infection in European badgers (Meles meles). 2. We investigated the relationship between badger abundance and M. bovis prevalence, using data on 2696 adult badgers in 10 populations sampled at the start of the Randomized Badger Culling Trial. 3. M. bovis prevalence was consistently higher at low badger densities and in small social groups. M. bovis prevalence was also higher among badgers whose genetic profiles suggested that they had immigrated into their assigned social groups. 4. The association between high M. bovis prevalence and small badger group size appeared not to have been caused by previous small-scale culling in study areas, which had been suspended, on average, 5 years before the start of the current study. 5. The observed pattern of prevalence might occur through badgers in smaller groups interacting more frequently with members of neighbouring groups; detailed behavioural data are needed to test this hypothesis. Likewise, longitudinal data are needed to determine whether the size of infected groups might be suppressed by disease-related mortality. 6. Although M. bovis prevalence was lower at high population densities, the absolute number of infected badgers was higher. However, this does not necessarily mean that the risk of M. bovis transmission to cattle is highest at high badger densities, since transmission risk depends on badger behaviour as well as on badger density.
Berg, Stefan; Garcia-Pelayo, M. Carmen; Müller, Borna; Hailu, Elena; Asiimwe, Benon; Kremer, Kristin; Dale, James; Boniotti, M. Beatrice; Rodriguez, Sabrina; Hilty, Markus; Rigouts, Leen; Firdessa, Rebuma; Machado, Adelina; Mucavele, Custodia; Ngandolo, Bongo Nare Richard; Bruchfeld, Judith; Boschiroli, Laura; Müller, Annélle; Sahraoui, Naima; Pacciarini, Maria; Cadmus, Simeon; Joloba, Moses; van Soolingen, Dick; Michel, Anita L.; Djønne, Berit; Aranaz, Alicia; Zinsstag, Jakob; van Helden, Paul; Portaels, Françoise; Kazwala, Rudovick; Källenius, Gunilla; Hewinson, R. Glyn; Aseffa, Abraham; Gordon, Stephen V.; Smith, Noel H.
We have identified a clonal complex of Mycobacterium bovis isolated at high frequency from cattle in Uganda, Burundi, Tanzania, and Ethiopia. We have named this related group of M. bovis strains the African 2 (Af2) clonal complex of M. bovis. Af2 strains are defined by a specific chromosomal deletion (RDAf2) and can be identified by the absence of spacers 3 to 7 in their spoligotype patterns. Deletion analysis of M. bovis isolates from Algeria, Mali, Chad, Nigeria, Cameroon, South Africa, and Mozambique did not identify any strains of the Af2 clonal complex, suggesting that this clonal complex of M. bovis is localized in East Africa. The specific spoligotype pattern of the Af2 clonal complex was rarely identified among isolates from outside Africa, and the few isolates that were found and tested were intact at the RDAf2 locus. We conclude that the Af2 clonal complex is localized to cattle in East Africa. We found that strains of the Af2 clonal complex of M. bovis have, in general, four or more copies of the insertion sequence IS6110, in contrast to the majority of M. bovis strains isolated from cattle, which are thought to carry only one or a few copies. PMID:21097608
CHENG, YAFEN; CHOU, CHUNG-HSI; TSAI, HSIANG-JUNG
The intracellular parasite Mycobacterium bovis (M. bovis) causes tuberculosis in cattle and humans. Understanding the interactions between M. bovis and host cells is essential in developing tools for the prevention, detection, and treatment of M. bovis infection. Gene expression profiles provide a large amount of information regarding the molecular mechanisms underlying these interactions. The present study analyzed changes in gene expression in bovine peripheral blood mononuclear cells (PBMCs) at 0, 4 and 24 h following exposure to M. bovis. Using bovine whole-genome microarrays, a total of 420 genes were identified that exhibited significant alterations in expression (≥2-fold). Significantly enriched genes were identified using the Kyoto Encyclopedia of Genes and Genomes database, of which the highest differentially expressed genes were associated with the immune system, signal transduction, endocytosis, cellular transport, inflammation, and apoptosis. Of the genes associated with the immune system, 84.85% displayed downregulation. These findings support the view that M. bovis inhibits signaling pathways of antimycobacterial host defense in bovine PBMCs. These in vitro data demonstrated that molecular alterations underlying the pathogenesis of tuberculosis begin early, during the initial 24 h following M. bovis infection. PMID:26668602
Labidi, A; Thoen, C O
Total DNA from two slowly-growing pathogenic mycobacterial species propagated in vitro was isolated, digested with each of 34 restriction endonucleases and analyzed by agarose gel electrophoresis. The most resolved patterns for M. tuberculosis (ATCC 27294) and for M. bovis (ATCC 19210) were obtained respectively using (BamHI, DraI, ClaI, EcoRI, EcoRV, HindIII, HpaI, SalI, SmaI, XbaI and XmaI). The patterns produced for these strains were reproducible and distinguishable from each other respectively using (HindIII, DraI, EcoRI, MboI, Sau3AI and AvaI). However, with several enzymes (SalI, AsuI, Sau96I, MspI and HpaII) the patterns for M. tuberculosis and M. bovis were similar. Evidence was obtained for the presence of dam and dcmI methylations in the DNA of each mycobacterial species respectively using (MboI, Sau3AI, EcoRII, BstNI, Sau96I and AsuI).
Manuel, Christopher A; Pugazhenthi, Umarani; Leszczynski, Jori K
Corynebacterium bovis causes an opportunistic infection of nude (Foxn1, nu/nu) mice, leading to nude mouse hyperkeratotic dermatitis (scaly skin disease). Enzootic in many nude mouse colonies, C. bovis spreads rapidly to naive nude mice, despite modern husbandry practices, and is very difficult to eradicate. To facilitate rapid detection in support of eradication efforts, we investigated a surveillance method based on quantitative real-time PCR (qPCR) evaluation of swabs collected from the horizontal exhaust manifold (HEM) of an IVC rack system. We first evaluated the efficacy of rack sanitation methods for removing C. bovis DNA from the HEM of racks housing endemic colonies of infected nude mice. Pressurized water used to flush the racks' air exhaust system followed by a standard rack-washer cycle was ineffective in eliminating C. bovis DNA. Only after autoclaving did all sanitized racks test negative for C. bovis DNA. We then measured the effects of stage of infection (early or established), cage density, and cage location on the rack on time-to-detection at the HEM. Stage of infection significantly affected time-to-detection, independent of cage location. Early infections required 7.3 ± 1.2 d whereas established infections required 1 ± 0 d for detection of C. bovis at the HEM. Cage density influenced the quantity of C. bovis DNA detected but not time-to-detection. The location of the cage on the rack affected the time-to-detection only during early C. bovis infections. We suggest that qPCR swabs of HEM are useful during the routine surveillance of nude mouse colonies for C. bovis infection.
Denis, Michel; Buddle, Bryce M
Bovine and possum macrophages were infected in vitro with a virulent strain of Mycobacterium bovis, and mycobacterial replication was measured in the infected macrophages cultured under a variety of conditions. Virulent M. bovis replicated substantially in alveolar possum macrophages as well as in bovine blood monocyte-derived macrophages. Addition of recombinant bovine interferon-gamma (IFN-gamma) with low concentrations of lipopolysaccharide (LPS) rendered bovine macrophages significantly more resistant to M. bovis replication. Disruption of iron levels in infected macrophages by addition of apotransferrin or bovine lactoferrin blocked replication of M. bovis in both bovine and possum macrophages. On the other hand, addition of exogenous iron, either in the form of iron citrate or iron-saturated transferrin, rendered macrophages of both species much more permissive for the replication of M. bovis. The impact of iron deprivation/loading on the mycobacteriostatic activity of cells was independent of nitric-oxide release, as well as independent of the generation of oxygen radical species in both possum and bovine macrophages. Exogenous iron was shown to reverse the ability of IFN-gamma/LPS pulsed bovine macrophages to restrict M. bovis replication. When autologous possum lymphocytes from animals vaccinated with M. bovis strain BCG were added to infected macrophages, they rendered the macrophages less permissive for virulent M. bovis replication. Loading the cells with iron prior to this macrophage-lymphocyte interaction, reversed this immune effect induced by sensitized cells. We conclude that, in two important animal species, intracellular iron level plays an important role in M. bovis replication in macrophages, irrespective of their activation status.
Manuel, Christopher A; Pugazhenthi, Umarani; Leszczynski, Jori K
Corynebacterium bovis causes an opportunistic infection of nude (Foxn1, nu/nu) mice, leading to nude mouse hyperkeratotic dermatitis (scaly skin disease). Enzootic in many nude mouse colonies, C. bovis spreads rapidly to naive nude mice, despite modern husbandry practices, and is very difficult to eradicate. To facilitate rapid detection in support of eradication efforts, we investigated a surveillance method based on quantitative real-time PCR (qPCR) evaluation of swabs collected from the horizontal exhaust manifold (HEM) of an IVC rack system. We first evaluated the efficacy of rack sanitation methods for removing C. bovis DNA from the HEM of racks housing endemic colonies of infected nude mice. Pressurized water used to flush the racks’ air exhaust system followed by a standard rack-washer cycle was ineffective in eliminating C. bovis DNA. Only after autoclaving did all sanitized racks test negative for C. bovis DNA. We then measured the effects of stage of infection (early or established), cage density, and cage location on the rack on time-to-detection at the HEM. Stage of infection significantly affected time-to-detection, independent of cage location. Early infections required 7.3 ± 1.2 d whereas established infections required 1 ± 0 d for detection of C. bovis at the HEM. Cage density influenced the quantity of C. bovis DNA detected but not time-to-detection. The location of the cage on the rack affected the time-to-detection only during early C. bovis infections. We suggest that qPCR swabs of HEM are useful during the routine surveillance of nude mouse colonies for C. bovis infection. PMID:26817981
Boothby, J T; Schore, C E; Jasper, D E; Osburn, B I; Thomas, C B
This study characterized the immune responses in four vaccinated and four control cows in response to vaccination and experimental intramammary inoculation with Mycoplasma bovis. Specific antibody responses occurred in serum and milk in response to vaccination and experimental infection. Lymphocytes from peripheral blood, but not from the mammary gland of vaccinated cows had increased responsiveness to mitogens. No lymphocytes tested were responsive to M. bovis antigen. Both vaccination and experimental infection resulted in skin test reactivity. These results imply that vaccination results in immune responses which may alter the course of experimental M. bovis mastitis, but may contribute to cellular inflammation. PMID:3167718
Kim, Tae-Hyoun; Kim, Dong-Su; Han, Ju-Hee; Chang, Seo-Na; Kim, Kyung-Sul; Seok, Seung-Hyeok; Kim, Dong-Jae; Park, Jong-Hwan; Park, Jae-Hak
Corynebacterium (C.) bovis infection in nude mice causes hyperkeratosis and weight loss and has been reported worldwide but not in Korea. In 2011, nude mice from an animal facility in Korea were found to have white flakes on their dorsal skin. Histopathological testing revealed that the mice had hyperkeratosis and Gram-positive bacteria were found in the skin. We identified isolated bacteria from the skin lesions as C. bovis using PCR and 16S rRNA sequencing. To the best of our knowledge, this is the first report of C. bovis infection in nude mice from Korea.
Matjila, P T; Penzhorn, B L; Leisewitz, A L; Bhoora, R; Barker, R
Canine babesiosis caused by Babesia gibsoni was diagnosed in a 3-month-old Pit-bull pup during a routine clinical examination. Diagnosis was confirmed by way of smear examination, PCR, Reverse Line Blot (RLB) and sequence analysis which showed 100% homology with B. gibsoni (Japan AB118032) and Babesia sp. (Oklahoma) (AF205636). Haematology showed moderate anaemia and severe thrombocytopenia. Treatment was initiated with diminazene aceturate (Berenil RTU) followed by 2 doses of imidocarb diproprionate (Forray-65) 3 days and 14 days later, respectively. Babesia gibsoni DNA was still detectable 2 weeks post-treatment on the PCR/RLB test. A 10-day course of combination drug therapy using atovaquone and azithromycin was initiated. Blood samples taken on Day 1 and Day 40 after completion of treatment were negative for B. gibsoni DNA on PCR/RLB test. The implications of a possible introduction of B. gibsoni into South Africa are discussed.
Forgrave, R; Donaghy, J A; Fisher, A; Rowe, M T
Reports have highlighted the absence of contemporary peer reviewed publications pertaining to Mycobacterium bovis culture from raw milk and cheese. By replicating traditional methods, cheese-making methodology and equipment were devised to produce Cheddar (n = 6) and Caerphilly (n = 3) artificially contaminated with M. bovis (three genotypes) under stringent laboratory-containment guidelines for handling hazardous microbiological material. Middlebrook 7H11, modified for M. bovis isolation, was assessed for capacity to enumerate M. bovis despite changing cheese microflora and prolonged M. bovis exposure to the cheese matrix using maturing cheese test portions (n = 63; up to 16 weeks). Malachite green (MG) containing media isolated M. bovis at significantly (P < 0·05) lower levels than unmodified Middlebrook 7H11 agar despite MG being a common adjunct of Middlebrook 7H11 agar modified for M. bovis growth. Subsequently, a selective MG-free Middlebrook 7H11 agar modified using haemolysed red cells and calf serum was demonstrated as the best performing (P < 0·05) medium for recovery of M. bovis from typical UK cheese types, Cheddar and Caerphilly. Significance and impact of the study: Following increased M. bovis infection of UK cattle, the risk posed to consumers from consumption of unpasteurized milk and dairy products has changed. Furthermore, published methods for the culture and molecular detection of M. bovis in raw milk products are limited. Cheese-making protocols and M. bovis culture media reported here provide tools for further investigation of M. bovis survival during all stages of cheese manufacture and could inform future assessment of the risk to consumers from M. bovis contamination of unpasteurized dairy products.
HIGA, Yumiko; UEMURA, Ryoko; YAMAZAKI, Wataru; GOTO, Shinya; GOTO, Yoshitaka; SUEYOSHI, Masuo
We improved a loop-mediated isothermal amplification (LAMP) assay permitting sensitive and rapid Mycoplasma bovis detection. A total of 55 bacterial strains were examined in this study, including 33 M. bovis strains, 14 non-M. bovis mycoplasmas and eight non-mycoplasma bacterial strains. M. bovis was successfully detected by the LAMP assay within 60 min without cross-reaction to any other bacteria. Furthermore, a total of 135 nasal swab samples were tested directly using our LAMP assays, the previously reported LAMP assay, conventional PCR assay without pre-culture and comparing standard culture methods. The improved LAMP assay showed sensitivity and specificity of 97.2% and 90.9%, respectively (with a kappa coefficient of 0.8231), and the sensitivity of our revised LAMP assay was increased compared to existing methods. PMID:27109067
Loutet, Miranda G.; O’Connor, Catherine; Kearns, Cathriona; Smith, Robert M.M.; Lalor, Maeve K.; Thomas, H. Lucy; Abubakar, Ibrahim; Zenner, Dominik
Despite control efforts, Mycobacterium bovis incidence among cattle remains high in parts of England, Wales, and Northern Ireland, attracting political and public health interest in potential spread from animals to humans. To determine incidence among humans and to identify associated factors, we conducted a retrospective cohort analysis of human M. bovis cases in England, Wales, and Northern Ireland during 2002–2014. We identified 357 cases and observed increased annual case numbers (from 17 to 35) and rates. Most patients were >65 years of age and born in the United Kingdom. The median age of UK-born patients decreased over time. For 74% of patients, exposure to risk factors accounting for M. bovis acquisition, most frequently consumption of unpasteurized milk, was known. Despite the small increase in case numbers and reduction in patient age, M. bovis infection of humans in England, Wales, and Northern Ireland remains rare. PMID:28220748
Pérez-Lago, Laura; Navarro, Yurena; García-de-Viedma, Darío
Mycobacterium bovis is both the causative agent of bovine tuberculosis (TB) and a zoonotic pathogen. In humans, considerably fewer cases of TB are caused by M. bovis than M. tuberculosis; nevertheless, diagnostic limitations mean that currently available data on prevalence grossly underestimate the true dimension of the problem. The routes of transmission from animals to humans are well known and include direct exposure to infected animals or consumption of contaminated animal products. Application of fingerprinting tools facilitates analysis of the molecular epidemiology of M. bovis in animal-to-human and human-to-human transmission. Apart from cattle and M. bovis, other animal species and members within the M. tuberculosis complex can contribute to the zoonosis. Improvements in diagnostic techniques, application of more advanced discriminatory genotyping tools, and collaboration between veterinary and human health care researchers are key to our understanding of this zoonosis.
Mycobacterium bovis isolates carry restricted allelic variation yet exhibit a range of disease phenotypes and host preferences. Conventional genotyping methods target small hyper-variable regions of their genome and provide anonymous biallelic information insufficient to develop phylogeny. To resolv...
Background: Bovine tuberculosis remains one of the most damaging zoonotic diseases. A critical need exists for rapid and inexpensive diagnostics capable of detecting and differentiating M. bovis infection from other pathogenic and environmental mycobacteria at multiple surveillance levels. Method...
Davidson, Jennifer A; Loutet, Miranda G; O'Connor, Catherine; Kearns, Cathriona; Smith, Robert M M; Lalor, Maeve K; Thomas, H Lucy; Abubakar, Ibrahim; Zenner, Dominik
Despite control efforts, Mycobacterium bovis incidence among cattle remains high in parts of England, Wales, and Northern Ireland, attracting political and public health interest in potential spread from animals to humans. To determine incidence among humans and to identify associated factors, we conducted a retrospective cohort analysis of human M. bovis cases in England, Wales, and Northern Ireland during 2002-2014. We identified 357 cases and observed increased annual case numbers (from 17 to 35) and rates. Most patients were >65 years of age and born in the United Kingdom. The median age of UK-born patients decreased over time. For 74% of patients, exposure to risk factors accounting for M. bovis acquisition, most frequently consumption of unpasteurized milk, was known. Despite the small increase in case numbers and reduction in patient age, M. bovis infection of humans in England, Wales, and Northern Ireland remains rare.
Rodríguez, Juan Germán; Fissanoti, Juan Carlos; Del Portillo, Patricia; Patarroyo, Manuel Elkin; Romano, María Isabel; Cataldi, Angel
The presence of a 500-bp fragment which amplifies a region from the genome of Mycobacterium bovis (J. G. Rodriguez, G. A. Meija, P. Del Portillo, M. E. Patarroyo, and L. A. Murillo, Microbiology 141:2131–2138, 1995) was evaluated by carrying out PCR on 121 M. bovis isolates. The M. bovis strains, previously characterized by culture and biochemical tests, were isolated from cattle in different regions of Argentina, Mexico, and Colombia. Four additional strains isolated from sea lions that belong to the M. tuberculosis complex were also included in the study. All of the isolates tested were PCR positive, rendering the expected 500-bp band and giving a correlation of 100% with previous microbiological characterization. Southern blot analysis revealed a common band of 1,800 bp and a polymorphic high-molecular-mass hybridization pattern. The results show that this assay may be useful for diagnosis and identification of M. bovis in cattle. PMID:10364607
Pathogenic mycobacteria of the Mycobacterium tuberculosis complex such as Mycobacterium bovis, induce a characteristic lesion known as a granulomas. Granulomas represent a specific host response to chronic antigenic stimuli, such as foreign bodies, certain bacterial components, or persistent pathoge...
Chatterjee, D; Bozic, C M; Knisley, C; Cho, S N; Brennan, P J
The glycolipid that characterizes the majority of isolates of Mycobacterium bovis and that has come to be known as M. bovis-identifying lipid is the phenolic glycolipid mycoside B described in the literature by others. However, when mycoside B obtained from M. bovis BCG, field isolates, and infected tissues was examined in detail, it was shown to be different from that described in the literature in some important respects. In particular, the glycosyl substituent is 2-O-methyl-alpha-L-rhamnopyranose rather than 2-O-methyl-beta-D-rhamnopyranose. With this information, a seroreactive neoglycoprotein (neoantigen) containing the 2-O-methyl-alpha-L-rhamnopyranosyl substituent suitable for the serodiagnosis of bovine tuberculosis was synthesized. M. bovis also contains other minor seroreactive phenolic glycolipids, one of which is a deacylated form of mycoside B and another of which contains an alpha-L-rhamnopyranosyl unit rather than 2-O-methyl-alpha-L-rhamnopyranose. Images PMID:2643563
Garrido, Joseba M.; Sevilla, Iker A.; Beltrán-Beck, Beatriz; Minguijón, Esmeralda; Ballesteros, Cristina; Galindo, Ruth C.; Boadella, Mariana; Lyashchenko, Konstantin P.; Romero, Beatriz; Geijo, Maria Victoria; Ruiz-Fons, Francisco; Aranaz, Alicia; Juste, Ramón A.; Vicente, Joaquín; de la Fuente, José; Gortázar, Christian
Tuberculosis (TB) caused by Mycobacterium bovis and closely related members of the Mycobacterium tuberculosis complex continues to affect humans and animals worldwide and its control requires vaccination of wildlife reservoir species such as Eurasian wild boar (Sus scrofa). Vaccination efforts for TB control in wildlife have been based primarily on oral live BCG formulations. However, this is the first report of the use of oral inactivated vaccines for controlling TB in wildlife. In this study, four groups of 5 wild boar each were vaccinated with inactivated M. bovis by the oral and intramuscular routes, vaccinated with oral BCG or left unvaccinated as controls. All groups were later challenged with a field strain of M. bovis. The results of the IFN-gamma response, serum antibody levels, M. bovis culture, TB lesion scores, and the expression of C3 and MUT genes were compared between these four groups. The results suggested that vaccination with heat-inactivated M. bovis or BCG protect wild boar from TB. These results also encouraged testing combinations of BCG and inactivated M. bovis to vaccinate wild boar against TB. Vaccine formulations using heat-inactivated M. bovis for TB control in wildlife would have the advantage of being environmentally safe and more stable under field conditions when compared to live BCG vaccines. The antibody response and MUT expression levels can help differentiating between vaccinated and infected wild boar and as correlates of protective response in vaccinated animals. These results suggest that vaccine studies in free-living wild boar are now possible to reveal the full potential of protecting against TB using oral M. bovis inactivated and BCG vaccines. PMID:21935486
Manuel, Christopher A; Pugazhenthi, Umarani; Spiegel, Shannon P; Leszczynski, Jori K
Rodent health-monitoring programs based on sampling an IVC system's exhaust air dust (EAD) has enhanced and even replaced traditional sentinels for some rodent pathogens. EAD testing by qPCR assay is an optimal surveillance method for the rapid detection of Corynebacterium bovis-infected immunodeficient mice. Here we demonstrate that an active EAD surveillance program for C. bovis can be used to maintain nude mice C. bovis-free after the transition from historically enzootically infected colonies. During 3 events over 3 y, rapid detection of infection, elimination of infected mice, aggressive quarantine measures, and local decontamination prevented the spread of C. bovis within 2 barrier rooms. In total, 4 cages of infected nude mice were identified and removed, preventing the spread of infection to 469 other cages of immunodeficient mice. In addition, we present data regarding a refinement to EAD testing which enables row-specific surveillance of an IVC rack. This technique systemically decreases the amount of testing required to locate an individually infected cage. Due to our ability to rapidly detect and localize an infected cage, we were able to investigate the route of C. bovis introduction into our barrier rooms. Our epidemiologic investigation suggested that the transmission of C. bovis occurred through contaminated, cryopreserved, patient-derived xenograft tumor tissue. This previously unknown source of C. bovis can infect mice used to propagate these tumors. Together, these data demonstrate that a remediation program that combines rapid detection, test-and-cull, and local decontamination under quarantine conditions can eliminate C. bovis from a mouse colony.
Carvalho, Ricardo César Tavares; Vasconcellos, Sidra Ezidio Gonçalves; Issa, Marina de Azevedo; Soares Filho, Paulo Martins; Mota, Pedro Moacyr Pinto Coelho; de Araújo, Flábio Ribeiro; Carvalho, Ana Carolina da Silva; Gomes, Harrison Magdinier; Suffys, Philip Noel; Paschoalin, Vânia Margaret Flosi
Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB), the pathogen responsible for serious economic impact on the livestock sector. In order to obtain data on isolated M. bovis strains and assist in the control and eradication program for BTB, a cross sectional descriptive molecular epidemiology study in the Brazilian Midwest was conducted. Through spoligotyping and 24-loci MIRU-VNTR methods, 37 clinical isolates of M. bovis circulating in the region were analyzed, 10 isolated from the state of Mato Grosso, 12 from the state of Mato Grosso do Sul and 15 from the state of Goiás. The spoligotyping analysis identified 10 distinct M. bovis profiles (SB0121 n = 14, SB0295 n = 6, SB0140 n = 6, SB0881 n = 3, SB1144 n = 2, SB1145 n = 2, SB0134 n = 1, SB1050 n = 1, SB1055 n = 1, SB1136 n = 1) grouped in six clusters and four orphan patterns. The MIRU-VNTR 24-loci grouped the same isolates in six clusters and 22 unique orphan patterns, showing higher discriminatory power than spoligotyping. When associating the results of both techniques, the isolates were grouped in five clusters and 24 unique M. bovis profiles. Among the 24-loci MIRU-VNTR evaluated, two, ETR-A and QUB 11b loci, showed high discriminatory ability (h = ≥ 0.50), while MIRU 16, MIRU 27, ETR-B, ETR-C, Mtub21 and QUB 26 loci showed moderate ability (h = 0.33 or h = 0.49) and were the most effective in evaluating the genotypic similarities among the clinical M. bovis isolate samples. Herein, the 29 patterns found amongst the 37 isolates of M. bovis circulating in the Brazilian Midwest can be due to the animal movement between regions, municipalities and farms, thus causing the spread of various M. bovis strains in herds from Midwest Brazil. PMID:27631383
Rocha, V.C.M.; Corrêa, S.H.R.; Oliveira, E.M.D.; C.A.R., Rodriguez; Fedullo, J.D.; Matrone, M.; Setzer, A.; Ikuta, C.Y.; Vejarano, M.P.; Figueiredo, S.M.; Ferreira Neto, J.S.
Two waterbucks from São Paulo Zoo Foundation exhibited respiratory symptoms in July 2004. After euthanasia, granulommas in lungs and mediastinic lymph nodes were observed. Acid-fast bacilli isolated were identified as Mycobacterium bovis spoligotype SB0121 by PRA and spoligotyping. They were born and kept in the same enclosure with the same group, without any contact to other species housed in the zoo. This is the first detailed description of M. bovis infection in Kobus ellipsiprymnus. PMID:24031687
Chapman, Rosamund; Chege, Gerald; Shephard, Enid; Stutz, Helen; Williamson, Anna-Lise
HIV-1 has resulted in a devastating AIDS pandemic. An effective HIV/AIDS vaccine that can be used to either, prevent HIV infection, control infection or prevent progression of the disease to AIDS is needed. In this review we discuss the use of Mycobacterium bovis BCG, the tuberculosis vaccine, as a vaccine vector for an HIV vaccine. Numerous features make BCG an attractive vehicle to deliver HIV antigens. It has a good safety profile, elicits long-lasting cellular immune responses and in addition manufacturing costs are affordable, a necessary consideration for developing countries. In this review we discuss the numerous factors that influence generation of a genetically stable recombinant BCG vaccine for HIV. PMID:20353397
Long, R; Nobert, E; Chomyc, S; van Embden, J; McNamee, C; Duran, R R; Talbot, J; Fanning, A
Globally, the proportion of all cases of tuberculosis (TB) caused by drug-resistant strains is increasing. We report the case of a Canadian citizen who acquired a highly drug-resistant strain of Mycobacterium bovis while visiting a relative with AIDS-related tuberculosis in Spain. The origin of the strain was traced using spoligotyping, a polymerase chain reaction (PCR)-based fingerprint technology, and the European DNA database. The level of primary drug resistance-all five first-line drugs and 19 of 21 second-line drugs-in this case was unprecedented in Canada. Isolation of this strain from a Canadian citizen represents the first report of its appearance in this hemisphere. The infection was contained and combined medical-surgical treatment delivered.
Fine, Amanda E.; Bolin, Carole A.; Gardiner, Joseph C.; Kaneene, John B.
Reisolation of Mycobacterium bovis from inoculated substrates was used to follow the persistence of viable M. bovis bacteria exposed to natural weather conditions over a 12-month period. Environmental factors were recorded continuously, and factors affecting M. bovis persistence (i.e., temperature, season, and substrate) were studied using survival analysis and Cox's proportional hazards regression. Persistence of M. bovis in the environment was significantly shorter in the spring/summer season, characterized by the highest average daily temperatures over the 12-month period. M. bovis persisted up to 88 days in soil, 58 days in water and hay, and 43 days on corn. These studies demonstrate that M. bovis bacteria persist long enough to represent a risk of exposure for cattle and/or wildlife and strengthen evidence that suggests cattle farm biosecurity and efforts to eliminate supplemental feeding of white-tailed deer will decrease the risk of bovine TB transmission among and between cattle and deer populations. PMID:21547222
Boldbaatar, Damdinsuren; Battsetseg, Badgar; Matsuo, Tomohide; Hatta, Takeshi; Umemiya-Shirafuji, Rika; Xuan, Xuenan; Fujisaki, Kozo
A cDNA encoding the vitellogenin receptor of the ixodid tick, Haemaphysalis longicornis Neumann (HlVgR) was cloned and characterized. The full-length cDNA is 5631 bp, including an intact ORF encoding an expected protein with 1782 amino acids. The deduced amino acid sequence of the HlVgR cDNA revealed two ligand-binding domains with four class A cysteine-rich repeats in the first domain and eight in the second domain similar to those of insect VgRs. The immunoblot analysis detected approximately 197 kDa protein in both tick ovary and egg. The developmental expression profile demonstrated that HlVgR mRNA exists throughout the ovarian development, and the transcriptional level is especially high in the previtellogenic period. Immuno electron microscopy analysis demonstrated that the localization of HlVgR is detected on the external surface of oocyte plasma membrane. RNAi showed that eggs of HlVgR dsRNA-injected adult ticks had not developed into fully mature oocytes and laid abnormal eggs. The Babesia parasite DNA was not detected in the eggs of HlVgR dsRNA-injected tick that fed on Babesia gibsoni infected dog, whereas it was detected in the eggs of PBS-injected ticks and noninjected ticks. Expression of HlVgR was increased by the vitellogenic hormone 20-hydroxyecdysone. These results indicate that HlVgR, which is produced by the developing oocytes, is essential for Vg uptake, egg development in the H. longicornis tick, and transovarial transmission of Babesia parasites.
El-Dakhly, Khaled Mohamed; Goto, Minami; Noishiki, Kaori; El-Nahass, El-Shaymaa; Sakai, Hiroki; Yanai, Tokuma; Takashima, Yasuhiro
Canine babesiosis constitutes a major global veterinary medical problem caused by tick-borne hemoparasites Babesia gibsoni and Babesia canis. Babesia gibsoni induces more severe clinical signs and is mainly transmitted by the ixodid Haemaphysalis longicornis. In Japan, B. gibsoni is primarily found in the western districts, with few records in the eastern parts. The aim of the current investigation was to evaluate distribution patterns of B. gibsoni infection in 9 Japanese islands and peninsulas using direct microscopy and PCR. Therefore, 196 hunting dogs were randomly sampled during the period from March to September 2011. Ages and sexes of dogs were identified. Direct microscopy of Giemsa-stained blood smear revealed pear-shaped piroplasms of B. gibsoni in 3 (1.6%) dogs. PCR was done initially with the universal primer set (B18S-F and B18S-R) amplifying the 1,665-bp portion of the 18S rRNA gene, followed by the specific primer set (Bg18F1 and Bg18R2) amplifying 2,363-bp fragments of the same gene. Accordingly, 84 (42.9%) and 8 (4.1%) dogs were positive, respectively. The current investigation shows that canine babesiosis was recorded in all islands except for Sado Island, Atsumi Peninsula, and Tanegashima Island. The highest infection rate was detected in the main island of Okinawa, while the lowest was on Ishigaki Island. Both sexes were non-significantly infected. However, the diversity of infection in islands was significantly different (P < 0.05). Although B. gibsoni has been previously found in western and eastern Japan, the present work highlights the prevalence of infection in many Japanese districts, including islands and peninsulas, giving realistic data that can facilitate treatment and control.
Azmi, Kifaya; Ereqat, Suheir; Nasereddin, Abedelmajeed; Al-Jawabreh, Amer; Baneth, Gad; Abdeen, Ziad
Ixodid ticks transmit various infectious agents that cause disease in humans and livestock worldwide. A cross-sectional survey on the presence of protozoan pathogens in ticks was carried out to assess the impact of tick-borne protozoa on domestic animals in Palestine. Ticks were collected from herds with sheep, goats and dogs in different geographic districts and their species were determined using morphological keys. The presence of piroplasms and Hepatozoon spp. was determined by PCR amplification of a 460-540bp fragment of the 18S rRNA gene followed by RFLP or DNA sequencing. A PCR-RFLP method based on the 18S rRNA was used in order to detect and to identify Hepatozoon, Babesia and Theileria spp. A total of 516 ticks were collected from animals in six Palestinian localities. Five tick species were found: Rhipicephalus sanguineus sensu lato, Rhipicephalus turanicus, Rhipicephalus bursa, Haemaphysalis parva and Haemaphysalis adleri. PCR-based analyses of the ticks revealed Theileria ovis (5.4%), Hepatozoon canis (4.3%), Babesia ovis (0.6%), and Babesia vogeli (0.4%). Theileria ovis was significantly associated with ticks from sheep and with R. turanicus ticks (p<0.01). H. canis was detected only in R. sanguineus s.l. and was significantly associated with ticks from dogs (p<0.01). To our knowledge, this is the first report describing the presence of these pathogens in ticks collected from Palestine. Communicating these findings with health and veterinary professionals will increase their awareness, and contribute to improved diagnosis and treatment of tick-borne diseases.
Corner, Leigh A L; O'Meara, D; Costello, E; Lesellier, S; Gormley, E
Populations of Eurasian badgers (Meles meles) with tuberculosis (Mycobacterium bovis infection) are a significant reservoir of infection for cattle in Ireland and the United Kingdom. In this study the distribution of infection, histological lesions and gross lesions was determined in a sample of 132 culled badgers from naturally-infected wild populations. Badgers were culled when an epidemiological investigation following a tuberculosis breakdown in a cattle herd implicated badgers as the probable source of infection. The definition of tuberculosis infection was based on the isolation of M. bovis from tissues or clinical samples. An accurate diagnosis of infection was achieved by culturing a wide range of lymph nodes (LN) and organ tissues (mean 32.1) and clinical samples (faeces and urine) from each badger. Infection was detected in 57/132 badgers (43.2%). Histological lesions consistent with tuberculosis were seen in 39/57 (68.4%) culture-positive and 7/75 (9.3%) culture-negative animals. Gross lesions were seen in only 30/57 (52.6%) infected badgers, leaving a high proportion (47.4%) of infected animals with latent infection (no grossly visible lesions). The most frequently infected tissues were the lungs and axillary LN, followed by the deep cervical LN, parotid LN and tracheobronchial LN. The data support the hypotheses that in badgers there are only two significant routes of infection, namely, the lower respiratory tract and bite wounds, and that badgers are very susceptible to infection but resistant to the development and progression of the disease. At all levels of disease severity, infection was found in widely dispersed anatomical locations suggesting that there is early dissemination of infection in the period preceding the development of active immunity.
de Azevedo Issa, Marina; Martins Soares Filho, Paulo; Fonseca Júnior, Antônio Augusto; Arrais Hodon, Mikael; Cristian Dos Santos, Lílian; Karlisson Pimenta Dos Reis, Jenner; Cerqueira Leite, Rômulo
For the definitive diagnosis of bovine tuberculosis, isolation of the etiologic agent is required. However, there is no consensus on the best methodology for isolation of Mycobacterium bovis in Brazil. This study evaluated the most used decontaminants and culture media in the country, in order to identify the best combination for the Brazilian samples. Three decontaminants - 2% sodium hydroxide (w/v), 0.75% hexadecylpiridinium chloride (w/v) and 5% sulphuric acid (v/v) and four culture media - 7H11 Middlebrook with additives and OADC supplement "A" (7H11 A), the same media with another supplement trademark (7H11 B), tuberculosis blood agar (B83) and Stonebrink's medium were compared. Regarding the isolation, there were no significant differences between the decontaminants and media combinations, except 7H11A combined to any decontaminant. However, the mean colonies score was significantly greater when the samples were decontaminated with 5% sulphuric acid and inoculated in 7H11 B or SB, without significant difference between them, although colonies appeared earlier on 7H11B than on SB. The trademark of OADC supplement influenced the isolation rate and the number of isolated colonies in Middlebrook 7H11. An incubation time of four weeks was required to detect all positive samples in 7H11 B after decontamination with 5% sulphuric acid but there was an increase in the number of colonies until the sixth week of incubation. Overall, the best strategy for the primary isolation of M. bovis from Brazilian samples was the decontamination with 5% sulphuric acid (final concentration) and inoculation in Middlebrook 7H11 medium formulated with OADC supplement "B".
Wan, Tien-Chun; Cheng, Fu-Yuan; Liu, Yu-Tse; Lin, Liang-Chuan; Sakata, Ryoichi
The purpose of the study was to investigate bioactive compounds of in vitro cultured Calculus Suis and natural Calculus Bovis obtained as valuable by-products from animals used for meat production. The results showed that the components of natural Calculus Bovis were rich in bilirubin and biliverdin and had higher content of essential amino acids. The major amino acids of in vitro cultured Calculus Suis were identified as glycine, alanine, glutamic acid and aspartic acid, and those for natural Calculus Bovis were found to be glutamic acid, aspartic acid, proline, and arginine. The methionine and cysteine contents of precursors for glutathione in natural Calculus Bovis were significantly higher than those of in vitro cultured Calculus Suis. The mineral contents of zinc, iron and manganese of natural Calculus Bovis were significantly higher than those of in vitro cultured Calculus Suis. The major bile acids in both products were cholic acid and dehydrocholic acid, respectively. The chenodeoxycholic and ursodeoxycholic acid content of in vitro cultured Calculus Suis was significantly higher than that of natural Calculus Bovis.
Lysnyansky, Inna; Freed, Mor; Rosales, Ruben S; Mikula, Inna; Khateb, Nihaya; Gerchman, Irena; van Straten, Michael; Levisohn, Sharon
The prevalence of Mycoplasma bovis in milk samples submitted to the Israeli National Service for Udder Health and Milk Quality was determined during the period 2004-2014 and the genetic pattern of the obtained isolates was assessed by multilocus sequence typing (MLST). Mycoplasma spp. were identified in 66 herds including M. bovis (n = 60), M. cottewii (n = 3), M. bovigenitalium (n = 2), M. alkalescens (n = 2) and M. yeatsii (n = 1). The proportion of M. bovis infected herds was relatively low (0-0.68%) in 2004-2007, increased to 3.77% during the 2008 outbreak, and ranged from 0.77 to 2.77% during the 2009-2014 period. Since 2008, about eight M. bovis positive dairy herds have been identified in Israel annually, with six of which on average being newly infected. MLST of 57 M. bovis isolates revealed that sequence type 10 was the dominant genotype identified in 60% of the herds. In conclusion, these data show that M. bovis is the main mycoplasmal mastitic pathogen in Israel.
Heuvelink, Annet; Reugebrink, Constance; Mars, Jet
Control of Mycoplasma bovis infections depends on good husbandry practices and antibiotic treatment. To allow more prudent use of antimicrobial drugs, there is a need for information on the susceptibility profile of this pathogen. The objective of the present study was to analyse the in vitro antimicrobial susceptibility of clinical M. bovis isolates in the Netherlands. The collection comprised 95 bovine isolates, originating from lungs (n=56), mastitis milk (n=27), and synovial fluid (n=12), collected between 2008 and 2014. Minimal inhibitory concentrations (MICs) were assessed by broth microdilution, both by using in-house prepared MIC plates and by using commercially available MIC plates. For each antimicrobial agent, the range of MIC results, the MIC50, and MIC90 values were calculated. M. bovis strains recently isolated in the Netherlands appeared to be characterized by relatively high MIC values for antimicrobial agents that, until now, have been recommended by the Dutch Association of Veterinarians for treating pneumonia caused by Mycoplasma species. Fluoroquinolones appeared to be the most efficacious in inhibiting M. bovis growth, followed by tulathromycin and oxytetracycline. The highest MIC values were obtained for erythromycin, tilmicosin, and tylosin. Future studies should be done on determining M. bovis specific clinical breakpoints, standardization of methods to determine MIC values as well as molecular studies on detection of antimicrobial resistance mechanisms of M. bovis isolates to develop PCR assays for determining resistance.
Mandal, M; Banerjee, P S; Garg, Rajat; Ram, Hira; Kundu, K; Kumar, Saroj; Kumar, G V P P S Ravi
Canine babesiosis is a vector borne disease caused by intra-erythrocytic apicomplexan parasites Babesia canis (large form) and Babesia gibsoni (small form), throughout the globe. Apart from few sporadic reports on the occurrence of B. gibsoni infection in dogs, no attempt has been made to characterize Babesia spp. of dogs in India. Fifteen canine blood samples, positive for small form of Babesia, collected from northern to eastern parts of India, were used for amplification of 18S rRNA gene (∼1665bp) of Babesia sp. and partial ITS1 region (∼254bp) of B. gibsoni Asian genotype. Cloning and sequencing of the amplified products of each sample was performed separately. Based on sequences and phylogenetic analysis of 18S rRNA and ITS1 sequences, 13 were considered to be B. gibsoni. These thirteen isolates shared high sequence identity with each other and with B. gibsoni Asian genotype. The other two isolates could not be assigned to any particular species because of the difference(s) in 18S rRNA sequence with B. gibsoni and closer identity with Babesiaoccultans and Babesiaorientalis. In the phylogenetic tree, all the isolates of B. gibsoni Asian genotype formed a separate major clade named as Babesia spp. sensu stricto clade with high bootstrap support. The two unnamed Babesia sp. (Malbazar and Ludhiana isolates) clustered close together with B. orientalis, Babesia sp. (Kashi 1 isolate) and B. occultans of bovines. It can be inferred from this study that 18S rRNA gene and ITS1 region are highly conserved among 13 B. gibsoni isolates from India. It is the maiden attempt of genetic characterization by sequencing of 18S rRNA gene and ITS1 region of B. gibsoni from India and is also the first record on the occurrence of an unknown Babesia sp. of dogs from south and south-east Asia.
Fecal Volatile Organic Ccompound Profiles from White-Tailed Deer (Odocoileus virginianus) as Indicators of Mycobacterium bovis Exposure or Mycobacterium bovis Bacille Calmette-Guerin (BCG) Vaccination
Stahl, Randal S.; Ellis, Christine K.; Nol, Pauline; Waters, W. Ray; Palmer, Mitchell; VerCauteren, Kurt C.
White-tailed deer (Odocoileus virginianus) serve as a reservoir for bovine tuberculosis, caused by Mycobacterium bovis, and can be a source of infection in cattle. Vaccination with M. bovis Bacille Calmette Guerin (BCG) is being considered for management of bovine tuberculosis in deer. Presently, no method exists to non-invasively monitor the presence of bovine tuberculosis in deer. In this study, volatile organic compound profiles of BCG-vaccinated and non-vaccinated deer, before and after experimental challenge with M. bovis strain 95–1315, were generated using solid phase microextraction fiber head-space sampling over suspended fecal pellets with analysis by gas chromatography/mass spectrometry. Chromatograms were processed using XCMS Online to characterize ion variation among treatment groups. The principal component scores resulting from significant (α = 0.05) ion responses were used to build linear discriminant analysis models. The sensitivity and specificity of these models were used to evaluate the feasibility of using this analytical approach to distinguish within group comparisons between pre- and post-M. bovis challenge: non-vaccinated male or female deer, BCG-vaccinated male deer, and the mixed gender non-vaccinated deer data. Seventeen compounds were identified in this analysis. The peak areas for these compounds were used to build a linear discriminant classification model based on principal component analysis scores to evaluate the feasibility of discriminating between fecal samples from M. bovis challenged deer, irrespective of vaccination status. The model best representing the data had a sensitivity of 78.6% and a specificity of 91.4%. The fecal head-space sampling approach presented in this pilot study provides a non-invasive method to discriminate between M. bovis challenged deer and BCG-vaccinated deer. Additionally, the technique may prove invaluable for BCG efficacy studies with free-ranging deer as well as for use as a non
Fecal Volatile Organic Ccompound Profiles from White-Tailed Deer (Odocoileus virginianus) as Indicators of Mycobacterium bovis Exposure or Mycobacterium bovis Bacille Calmette-Guerin (BCG) Vaccination.
Stahl, Randal S; Ellis, Christine K; Nol, Pauline; Waters, W Ray; Palmer, Mitchell; VerCauteren, Kurt C
White-tailed deer (Odocoileus virginianus) serve as a reservoir for bovine tuberculosis, caused by Mycobacterium bovis, and can be a source of infection in cattle. Vaccination with M. bovis Bacille Calmette Guerin (BCG) is being considered for management of bovine tuberculosis in deer. Presently, no method exists to non-invasively monitor the presence of bovine tuberculosis in deer. In this study, volatile organic compound profiles of BCG-vaccinated and non-vaccinated deer, before and after experimental challenge with M. bovis strain 95-1315, were generated using solid phase microextraction fiber head-space sampling over suspended fecal pellets with analysis by gas chromatography/mass spectrometry. Chromatograms were processed using XCMS Online to characterize ion variation among treatment groups. The principal component scores resulting from significant (α = 0.05) ion responses were used to build linear discriminant analysis models. The sensitivity and specificity of these models were used to evaluate the feasibility of using this analytical approach to distinguish within group comparisons between pre- and post-M. bovis challenge: non-vaccinated male or female deer, BCG-vaccinated male deer, and the mixed gender non-vaccinated deer data. Seventeen compounds were identified in this analysis. The peak areas for these compounds were used to build a linear discriminant classification model based on principal component analysis scores to evaluate the feasibility of discriminating between fecal samples from M. bovis challenged deer, irrespective of vaccination status. The model best representing the data had a sensitivity of 78.6% and a specificity of 91.4%. The fecal head-space sampling approach presented in this pilot study provides a non-invasive method to discriminate between M. bovis challenged deer and BCG-vaccinated deer. Additionally, the technique may prove invaluable for BCG efficacy studies with free-ranging deer as well as for use as a non
Wanzala, Sylvia I; Nakavuma, Jesca; Travis, Dominic A; Kia, Praiscillia; Ogwang, Sam; Sreevatsan, Srinand
Bovine tuberculosis (BTB), a zoonotic infection of cattle caused by Mycobacterium bovis, results in losses of $3 billion to the global agricultural industry and represents the fourth most important livestock disease worldwide. M. bovis as a source of human infection is likely underreported due to the culture medium conditions used to isolate the organism from sputum or other sample sources. We report here the draft genome sequences of M. bovis BZ 31150, isolated from a bronchial washing from a captive chimpanzee, and M. bovis B2 7505, isolated from a human sputum sample in Uganda.
Zaugg, J L; Lane, V M
We evaluated the efficacy of buparvaquone in eliminating Babesia equi of European origin in carrier horses and in experimentally infected splenectomized ponies. When administered at the rate of 2.5 mg/kg of body weight, IM, 4 times at 96-hour intervals, buparvaquone was effective in eliminating B equi carrier infection in 1 horse. Such results could not be repeated at the same dosage or at 3.5 or 5 mg/kg, IM. Buparvaquone given at the rate of 4 to 6 mg/kg IV and/or IM was therapeutically effective in 4 of 5 acute B equi infections in splenectomized ponies. The treated ponies became carriers.
Punyapornwithaya, V; Fox, L K; Hancock, D D; Gay, J M; Wenz, J R; Alldredge, J R
The objective was to determine the incidence and transmission of mycoplasma mastitis in the hospital pen in a dairy herd of 650 lactating cows after a hospital pen was established following an outbreak of this disease. Mycoplasma mastitis status was monitored for 3 months through repeated collection of milk samples from cows with clinical mastitis (CM) and from bulk tank milk. During the outbreak 13 cows were diagnosed with Mycoplasma bovis CM, 1 cow with Mycoplasma sp. mastitis and 8 cows showed signs of arthritis, 3 of which were confirmed as having M. bovis arthritis. M. bovis isolates from cows with CM, arthritis and bulk tank milk had indistinguishable chromosomal digest pattern fingerprints. Incidence rates of M. bovis CM cases in the milking and hospital pens were 0.01 and 1.7 cases per 100 cow-days at risk. Approximately 70% of cows with M. bovis CM became infected within 12 days of entering the hospital pen. Transmission of M. bovis in the hospital pen occurred as 3 episodes. Each episode corresponded to the introduction of a cow with M. bovis CM from a milking pen. Evidence indicates that cows with M. bovis CM from milking pens were the source of transmission of the disease in the hospital pen and thus their presence in the hospital pen appeared to be a risk factor for transmission of M. bovis mastitis in this single case study herd.
Cuesta, Isabel; González, Luis M.; Estrada, Karel; Grande, Ricardo; Zaballos, Ángel; Lobo, Cheryl A.; Barrera, Jorge
Babesia divergens causes significant morbidity and mortality in cattle and splenectomized or immunocompromised individuals. Here, we present a 10.7-Mb high-quality draft genome of this parasite close to chromosome resolution that will enable comparative genome analyses and synteny studies among related parasites. PMID:25395649
Background: Equine piroplasmosis caused by Theileria equi, Babesia caballi, or both, cause significant economic losses in the equine industry and remains uncontrolled in Egypt. Methods: T. equi and B. caballi infections were assessed in blood from 88 horses and 51 donkeys from different localities ...
Paulauskas, Algimantas; Radzijevskaja, Jana; Karvelienė, Birutė; Grigonis, Aidas; Aleksandravičienė, Asta; Zamokas, Gintaras; Babickaitė, Lina; Sabūnas, Vytautas; Petkevičius, Saulius
Canine babesiosis caused by Babesia canis is an emerging infectious disease in Europe. Although previously uncommon, canine babesiosis has become quite frequent in Lithuania during the past decade. In the last few years an increasing number of cases with a wide variety of clinical signs have been recorded throughout the country. In Lithuania the identification of the disease agent in veterinarian clinics is based on a microscopic analysis of size and morphology. To date, no data on the genetic characterization of Babesia species in dogs have been documented for Lithuania. A total of 123 blood samples from dogs showing clinical signs of babesiosis on the basis of veterinary examination were tested for the presence of babesial parasites. Babesia isolated from dogs were detected and characterized by nested-PCR and sequence analysis of a fragment of the 18S rRNA gene. Babesia parasites were detected in blood smears of 94 dogs (76.4%). The molecular analysis revealed the presence of B. canis in 108 dogs (87.8%). Two genotypes of B. canis were distinguished on the basis on two nucleotide (GA → AG) substitutions observed in 18S rRNA gene sequences. The results of the present study provide knowledge of the distribution of B. canis genotypes in dogs in Lithuania, and show the necessity to use a molecular analysis for an accurate diagnosis of canine babesiosis.
Oliveira, M C S; Oliveira-Sequeira, T C G; Regitano, L C A; Alencar, M M; Néo, T A; Silva, A M; Oliveira, H N
Babesia bigemina infections were investigated in four genetic groups of beef cattle and in Rhipicephalus (Boophilus) microplus engorged female ticks. Blood samples and engorged female ticks were collected from 15 cows and 15 calves from each of the following genetic groups: Nelore, Angus x Nelore, Canchim x Nelore, and Simmental x Nelore. Microscopic examination of blood smears and tick hemolymph revealed that merozoites of B. bigemina (6/60) as well as kinetes of Babesia spp. (9/549) were only detected in samples (blood and ticks, respectively) originated from calves. PCR-based methods using primers for specific detection of B. bigemina revealed 100% infection in both calves and cows, regardless the genetic group. Tick infection was detected by nested-PCR amplifications showing that the frequency of B. bigemina was higher (P<0.01) in female ticks collected from calves (134/549) than in those collected from cows (52/553). The frequency of B. bigemina was similar in ticks collected from animals, either cows or calves, of the four genetic groups (P>0.05).
Trotta, Michele; Nicetto, Martina; Fogliazza, Alessandro; Montarsi, Fabrizio; Caldin, Marco; Furlanello, Tommaso; Solano-Gallego, Laia
The aims of this study were to determine natural infections by Anaplasma phagocytophilum/Anaplasma platys, Bartonella henselae, Ehrlichia canis, Leishmania infantum, Rickettsia spp., Babesia spp., and Hepatozoon spp. by molecular methods in ticks (n=91) removed from dogs with clinical signs and laboratory abnormalities compatible with tick-borne diseases (n=22) living in Italy and to assess the distribution and species of ticks encountered. Ticks from dogs living in southern Italy were all identified as Rhipicephalus sanguineus (n=25), ticks from central Italy included Rh. sanguineus (n=8) and Ixodes ricinus (n=9), ticks from northern Italy included Rh. sanguineus (n=45), Dermacentor marginatus (n=3), and one I. ricinus. Leishmania infantum, Rickettsia spp., and Babesia canis were the only pathogens detected in 7 (8%), 4 (4%), and 2 (2%) out of 91 ticks, respectively. L. infantum was detected in I. ricinus from central Italy and in Rh. sanguineus from northern and central Italy. Rickettsia conorii and Ri. massiliae were detected in Rh. sanguineus ticks from central and southern Italy (Sicily), respectively. Bab. canis was detected in D. marginatus ticks from northern Italy.
Imre, Mirela; Farkas, Róbert; Ilie, Marius Stelian; Imre, Kálmán; Dărăbuş, Gheorghe
Blood samples from 49 symptomatic dogs from 5 western and north-western counties of Romania were screened using microscopic examination, polymerase-chain-reaction-restriction-fragment-length-polymorphism procedure (PCR-RFLP), and sequence analysis. Results of the microscopic evaluation of stained blood smears revealed 45 positive samples with the presence of large and small intraerythrocytic piroplasms in 35 and 10 samples, respectively. Babesia canis (35/49, 71.4%) and Babesia gibsoni (14/49, 28.6%) were identified and differentiated by PCR-RFLP targeting the 18S rRNA gene. Results of the sequence analysis of all B. gibsoni and 17 randomly selected B. canis PCR products confirmed the PCR-RFLP-diagnosed species. The distribution of B. gibsoni infection was positively associated (p<0.001) with fighting dog breeds including infection in 12 American Pit Bull Terriers and one American Staffordshire Terrier. This report is the first to present molecular evidence of the occurrence of B. gibsoni in Romania confirmed by sequencing.
Terao, Masashi; Akter, Shirin; Yasin, Md Golam; Nakao, Ryo; Kato, Hirotomo; Alam, Mohammad Zahangir; Katakura, Ken
Babesia gibsoni is a tick-borne hemoprotozoan parasite of dogs that often causes fever and hemolytic illness. Detection of B. gibsoni has been predominantly reported in Asian countries, including Japan, Korea, Taiwan, Malaysia, Bangladesh and India. The present study shows the first molecular characterization of B. gibsoni detected from dogs in Bangladesh. Blood samples were collected on FTA® Elute cards from 50 stray dogs in Mymensingh District in Bangladesh. DNA eluted from the cards was subjected to nested PCR for the 18S rRNA gene of Babesia species. Approximately 800bp PCR products were detected in 15 of 50 dogs (30%). Based on restriction fragment length polymorphism (RFLP) and direct sequencing of the PCR products, all parasite isolates were identified as B. gibsoni. Furthermore, the BgTRAP (B. gibsoni thrombospondin-related adhesive protein) gene fragments were detected in 13 of 15 18S rRNA gene PCR positive blood samples. Phylogenetic analysis of the BgTRAP gene revealed that B. gibsoni parasites in Bangladesh formed a cluster, which was genetically different from other Asian B. gibsoni isolates. In addition, tandem repeat analysis of the BgTRAP gene clearly showed considerable genetic variation among Bangladeshi isolates. These results suggested that B. gibsoni parasites in a different genetic clade are endemic in dogs in Bangladesh. Further studies are required to elucidate the origin, distribution, vector and pathogenesis of B. gibsoni parasites circulating in dogs in Bangladesh.
Renneker, S; Abdo, J; Bakheit, M A; Kullmann, B; Beyer, D; Ahmed, J; Seitzer, U
Infections of small ruminants with Anaplasma, Theileria and Babesia species are widely distributed in the old world and are of great economic impact. In Iraq, data on disease occurrence in sheep caused by above-mentioned infectious agents are scarce. This study provides information on various haemoparasitic agents infecting sheep in the Kurdistan Region, Iraq, using molecular diagnostic tools. Altogether, 195 samples originating from three governorates in the Kurdistan Region, namely Duhok, Erbil and Sulaimaniya, were analysed. The following pathogens were identified: Anaplasma ovis (62.6%), Theileria ovis (14.35%), T. lestoquardi (7.7%), T. uilenbergi (5.6%) and Babesia ovis (1.5%). T. uilenbergi is detected for the first time in Iraq. Coinfection of sheep with different pathogens could be observed in this study, and it was found that 45 of 195 (23%) of the samples contained more than one pathogen. Even triple-positive samples were identified in 3% of the investigated animals. In conclusion, we confirm the coinfection of sheep with various haemoparasitic pathogen species in the Kurdistan Region of Iraq. Further investigations are needed to reveal the epidemiology of the diseases, the respective tick vectors, and, in the case of coinfection, pathogens' interaction and possible cross-protection.
Erdélyi, Károly; Mezősi, László; Vladov, Sztojkov; Földvári, Gábor
Two adult male Eurasian grey wolves belonging to a group of 12 animals, kept in an open air 15,000-m(2) enclosure at the Bear Farm facility near Veresegyháza, Hungary, were found dead in September 2002. Another 2 wolves died during the same period, but laboratory examination of their carcasses was not possible. During necropsy both animals were found to be in a good body condition. Oral mucosa, conjunctiva, sclera, and subcutaneous tissues revealed severe jaundice. The liver, gall bladder, and spleen were enlarged. The kidneys were paler than normal, and petechial haemorrhages were also seen under their fascia. Small, round Babesia-like organisms, 1.5-2 μm in diameter, were demonstrated in large numbers in stained impression smears made from the spleens of both animals. PCR amplification and sequencing identified Babesia canis. There are very few reports on babesiosis in the grey wolf, and our findings draw attention to the potential threat posed by B. canis that will probably have to be taken into account in future ex situ and in situ wolf conservation efforts.
HABIBI, Gholamreza; ESMAEILNIA, Kasra; HABLOLVARID, Mohammad Hasan; AFSHARI, Asghar; ZAMEN, Mohsen; BOZORGI, Soghra
Background: Equine piroplasmosis (EP) is the cause of persistent tick-borne infection with no symptoms, but the most important problem of EP is due to the persistent carrier state. Carrier animals to Babesia (Theileria) equi (Laveran 1901) and B. caballi (Nuttall, 1910) infestation could be identified by extremely sensitive PCR-based method. The purpose of this study was to identify the causative agents of equine piroplasmosis based on molecular and microscopic assays in equids from Kurdistan Province, Iran. Methods: Thirty one horse and mule blood samples were used with history of living in Kurdistan Province of Iran. The blood specimens were utilized for T. equi and B. caballi DNA identification by PCR and Giemsa stained smears for microscopic observation. Results: The results clearly showed the presence of B. (Theileria) equi DNA in 30 of 31 blood samples (96.77%), but the microscopic examination revealed the 3 of 31 positive Babesia like organisms in the red blood cells (9.67%). Conclusion: The obtained results demonstrated the presence of hidden B. (Theileria) equi infection in horses with previous habitance in Kurdistan Province of Iran. The carrier animals became a main source of infection and can transmit the disease. Therefore, hidden infection might be considered as a health threatening and limiting factor in animals used in therapeutic antisera research and production centers. PMID:27095973
Li, Pei; Wang, Rui; Dong, Wenqi; Hu, Linlin; Zong, Bingbing; Zhang, Yanyan; Wang, Xiangru; Guo, Aizhen; Zhang, Anding; Xiang, Yaozu; Chen, Huanchun; Tan, Chen
Mycobacterium bovis (M. bovis), the most common pathogens of tuberculosis (TB), is virulent to human and cattle, and transmission between cattle and humans warrants reconsideration concerning food safety and public health. Recently, efforts have begun to analyze cellular proteomic responses induced by Mycobacterium tuberculosis (M. tb). However, the underlying mechanisms by which virulent M. bovis affects human hosts are not fully understood. For the present study, we utilized a global and comparative labeling strategy of isobaric tag for relative and absolute quantitation (iTRAQ) to assess proteomic changes in the human monocyte cell line (THP-1) using a vaccine strain and two virulent strains H37Rv and M. bovis. We measured 2,032 proteins, of which 61 were significantly differentially regulated. Ingenuity Pathway Analysis was employed to investigate the canonical pathways and functional networks involved in the infection. Several pathways, most notably the phagosome maturation pathway and TNF signaling pathway, were differentially affected by virulent strain treatment, including the key proteins CCL20 and ICAM1. Our qRT-PCR results were in accordance with those obtained from iTRAQ. The key enzyme MTHFD2, which is mainly involved in metabolism pathways, as well as LAMTOR2 might be effective upon M. bovis infection. String analysis also suggested that the vacuolar protein VPS26A interacted with TBC1D9B uniquely induced by M. bovis. In this study, we have first demonstrated the application of iTRAQ to compare human protein alterations induced by virulent M. bovis infections, thus providing a conceptual understanding of mycobacteria pathogenesis within the host as well as insight into preventing and controlling TB in human and animal hosts' transmission. PMID:28337427
ter Laak, E A; Noordergraaf, J H; Verschure, M H
The purpose of this study was to determine the susceptibility of various strains of Mycoplasma bovis, Mycoplasma dispar, and Ureaplasma diversum, which are prevalent causes of pneumonia in calves, to 16 antimicrobial agents in vitro. The MICs of the antimicrobial agents were determined by a serial broth dilution method for 16 field strains and the type strain of M. bovis, for 19 field strains and the type strain of M. dispar, and for 17 field strains of U. diversum. Final MICs for M. bovis and M. dispar were read after 7 days and final MICs for U. diversum after 1 to 2 days. All strains tested were susceptible to tylosin, kitasamycin, and tiamulin but were resistant to nifuroquine and streptomycin. Most strains of U. diversum were intermediately susceptible to oxytetracycline but fully susceptible to chlortetracycline; most strains of M. bovis and M. dispar, however, were resistant to both agents. Strains of M. dispar and U. diversum were susceptible to doxycycline and minocycline, but strains of M. bovis were only intermediately susceptible. Susceptibility or resistance to chloramphenicol, spiramycin, spectinomycin, lincomycin, or enrofloxacin depended on the species but was not equal for the three species. The type strains of M. bovis and M. dispar were more susceptible to various antimicrobial agents, including tetracyclines, than the field strains. This finding might indicate that M. bovis and M. dispar strains are becoming resistant to these agents. Antimicrobial agents that are effective in vitro against all three mycoplasma species can be considered for treating mycoplasma infections in pneumonic calves. Therefore, tylosin, kitasamycin, and tiamulin may be preferred over oxytetracycline and chlortetracycline. PMID:8452363
Herrera-Rodríguez, Sara E.; Gordiano-Hidalgo, María Alejandra; López-Rincón, Gonzálo; Bojorquez-Narváez, Luis; Padilla-Ramírez, Francisco Javier; Pereira-Suárez, Ana Laura; Flores-Valdez, Mario Alberto
Bovine tuberculosis (bTB) remains a problem on many dairy farms in Mexico, as well as a public health risk. We previously found a high frequency of Mycobacterium bovis DNA in colostrum from dairy cows using a nested PCR to detect mpb70. Since there are no reliable in vivo tests to determine the effectiveness of booster Mycobacterium bovis BCG vaccination against bTB, in this work we monitored M. bovis DNA in colostrum by using this nested PCR. In order to decrease the risk of adverse reactions in animals likely containing viable M. bovis, a single application of BCG and a subunit vaccine (EEP-1) formulated with M. bovis culture filtrate proteins (CFP) and a copolymer as the adjuvant was performed in tuberculin skin test-negative cattle (TST−), while TST reactor animals (TST+) received EEP-1 only. Booster immunization using EEP-1 was applied to both groups, 2 months after primary vaccination to whole herds and 12 months later to lactating cows. Colostrum samples were collected from 6 farms where the cows were vaccinated over a 12-month period postvaccination and, for comparison, from one control farm where the cows were not vaccinated with comparable bTB prevalence. We observed an inverse relationship between the frequency of M. bovis DNA detection and time postvaccination at the first (P < 0.001) and second (P < 0.0001) 6-month periods. Additionally, the concentration of gamma interferon (IFN-γ) was higher in mpb70 PCR-positive colostrum samples (P = 0.0003). These results suggest that M. bovis DNA frequency in colostrum could be a potentially useful biomarker for bTB vaccine efficacy on commercial dairy farms. PMID:23425597
Herrera-Rodríguez, Sara E; Gordiano-Hidalgo, María Alejandra; López-Rincón, Gonzálo; Bojorquez-Narváez, Luis; Padilla-Ramírez, Francisco Javier; Pereira-Suárez, Ana Laura; Flores-Valdez, Mario Alberto; Estrada-Chávez, Ciro
Bovine tuberculosis (bTB) remains a problem on many dairy farms in Mexico, as well as a public health risk. We previously found a high frequency of Mycobacterium bovis DNA in colostrum from dairy cows using a nested PCR to detect mpb70. Since there are no reliable in vivo tests to determine the effectiveness of booster Mycobacterium bovis BCG vaccination against bTB, in this work we monitored M. bovis DNA in colostrum by using this nested PCR. In order to decrease the risk of adverse reactions in animals likely containing viable M. bovis, a single application of BCG and a subunit vaccine (EEP-1) formulated with M. bovis culture filtrate proteins (CFP) and a copolymer as the adjuvant was performed in tuberculin skin test-negative cattle (TST(-)), while TST reactor animals (TST(+)) received EEP-1 only. Booster immunization using EEP-1 was applied to both groups, 2 months after primary vaccination to whole herds and 12 months later to lactating cows. Colostrum samples were collected from 6 farms where the cows were vaccinated over a 12-month period postvaccination and, for comparison, from one control farm where the cows were not vaccinated with comparable bTB prevalence. We observed an inverse relationship between the frequency of M. bovis DNA detection and time postvaccination at the first (P < 0.001) and second (P < 0.0001) 6-month periods. Additionally, the concentration of gamma interferon (IFN-γ) was higher in mpb70 PCR-positive colostrum samples (P = 0.0003). These results suggest that M. bovis DNA frequency in colostrum could be a potentially useful biomarker for bTB vaccine efficacy on commercial dairy farms.
Malama, Sydney; Johansen, Tone Bjordal; Muma, John Bwalya; Mwanza, Sydney; Djønne, Berit; Godfroid, Jacques
Bovine tuberculosis (BTB) is a chronic bacterial disease caused by Mycobacterium bovis. Infections due to M. bovis, which serves as a stable reservoir, can pose serious challenge to control and eradicate in both wildlife and livestock at the interface. This study aimed at isolating and characterizing M. bovis from Kafue lechwe (Kobus leche kafuensis) and black lechwe (Kobus leche smithemani) at the animal/human interface in Zambia. The samples with lesions compatible with BTB collected during the hunting seasons of 2009 and 2010 were cultured for isolation of mycobacteria using Stonebrink with pyruvate (BD Diagnostics, MD, USA) and Middlebrook 7H10 (BD Diagnostics) slants. Isolated mycobacteria were identified using IS6110 polymerase chain reaction and deletion analysis. Molecular characterization of the isolates was performed using spoligotyping and mycobacteria interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) with nine loci. Data was analyzed using BioNumerics software 6.1. Out of the 39 samples, acid fast bacilli were detected in 27 (69.2 %) based on smear microscopy. Seven isolates were found to belong to Mycobacterium tuberculosis complex, and all were identified as M. bovis based on deletion analysis. All seven isolates were identical on spoligotyping as belonging to the SB0120 (SIT 482). MIRU-VNTR differentiated the isolates into five different patterns. This study has confirmed that M. bovis circulates in the Kafue lechwe, and non-tuberculous mycobacteria were detected in the black lechwe in Zambia which represents a wildlife reservoir, with a potential to spillover to cattle and humans. Isolates of M. bovis from lechwe antelopes are much conserved as only one spoligotype was detected. The study has shown that three loci differentiated fairly well. This option is cheap and less laborious, and hence a better option in resource-strained country like Zambia. The study further showed that some of the loci recommended by the European
ter Laak, E A; Noordergraaf, J H; Verschure, M H
The purpose of this study was to determine the susceptibility of various strains of Mycoplasma bovis, Mycoplasma dispar, and Ureaplasma diversum, which are prevalent causes of pneumonia in calves, to 16 antimicrobial agents in vitro. The MICs of the antimicrobial agents were determined by a serial broth dilution method for 16 field strains and the type strain of M. bovis, for 19 field strains and the type strain of M. dispar, and for 17 field strains of U. diversum. Final MICs for M. bovis and M. dispar were read after 7 days and final MICs for U. diversum after 1 to 2 days. All strains tested were susceptible to tylosin, kitasamycin, and tiamulin but were resistant to nifuroquine and streptomycin. Most strains of U. diversum were intermediately susceptible to oxytetracycline but fully susceptible to chlortetracycline; most strains of M. bovis and M. dispar, however, were resistant to both agents. Strains of M. dispar and U. diversum were susceptible to doxycycline and minocycline, but strains of M. bovis were only intermediately susceptible. Susceptibility or resistance to chloramphenicol, spiramycin, spectinomycin, lincomycin, or enrofloxacin depended on the species but was not equal for the three species. The type strains of M. bovis and M. dispar were more susceptible to various antimicrobial agents, including tetracyclines, than the field strains. This finding might indicate that M. bovis and M. dispar strains are becoming resistant to these agents. Antimicrobial agents that are effective in vitro against all three mycoplasma species can be considered for treating mycoplasma infections in pneumonic calves. Therefore, tylosin, kitasamycin, and tiamulin may be preferred over oxytetracycline and chlortetracycline.
Zbinden, Christina; Pilo, Paola; Frey, Joachim; Bruckmaier, Rupert M; Wellnitz, Olga
Mycoplasma bovis is an emerging bacterial agent causing bovine mastitis. Although these cell wall-free bacteria lack classical virulence factors, they are able to activate the immune system of the host. However, effects on the bovine mammary immune system are not yet well characterized and detailed knowledge would improve the prevention and therapy of mycoplasmal mastitis. The aim of this study was to investigate the immunogenic effects of M. bovis on the mammary gland in an established primary bovine mammary epithelial cell (bMEC) culture system. Primary bMEC of four different cows were challenged with live and heat-inactivated M. bovis strain JF4278 isolated from acute bovine mastitis, as well as with the type strain PG45. The immune response was evaluated 6 and 24h after mycoplasmal challenge by measuring the relative mRNA expression of selected immune factors by quantitative PCR. M. bovis triggered an immune response in bMEC, reflected by the upregulation of tumor necrosis factor-α, interleukin(IL)-1β, IL-6, IL-8, lactoferrin, Toll-like receptor-2, RANTES, and serum amyloid A mRNA. Interestingly, this cellular reaction was only observed in response to live, but not to heat-inactivated M. bovis, in contrast to other bacterial pathogens of mastitis such as Staphylococcus aureus. This study provides evidence that bMEC exhibit a strong inflammatory reaction in response to live M. bovis. The lack of a cellular response to heat-inactivated M. bovis supports the current hypothesis that mycoplasmas activate the immune system through secreted secondary metabolites.
Yang, Yin-Shan; Murciano, Brice; Moubri, Karina; Cibrelus, Prisca; Schetters, Theo; Gorenflot, André; Delbecq, Stéphane; Roumestand, Christian
Babesiosis (formerly known as piroplasmosis) is a tick-borne disease caused by the intraerythrocytic development of protozoa parasites from the genus Babesia. Like Plasmodium falciparum, the agent of malaria, or Toxoplasma gondii, responsible for human toxoplasmosis, Babesia belongs to the Apicomplexa family. Babesia canis is the agent of the canine babesiosis in Europe. Clinical manifestations of this disease range from mild to severe and possibly lead to death by multiple organ failure. The identification and characterization of parasite surface proteins represent major goals, both for the understanding of the Apicomplexa invasion process and for the vaccine potential of such antigens. Indeed, we have already shown that Bd37, the major antigenic adhesion protein from Babesia divergens, the agent of bovine babesiosis, was able to induce complete protection against various parasite strains. The major merozoite surface antigens of Babesia canis have been described as a 28-kDa membrane protein family, anchored at the surface of the merozoite. Here, we demonstrate that Bc28.1, a major member of this multigenic family, is expressed at high levels at the surface of the merozoite. This protein is also found in the parasite in vitro culture supernatants, which are the basis of effective vaccines against canine babesiosis. We defined the erythrocyte binding function of Bc28.1 and determined its high resolution solution structure using NMR spectroscopy. Surprisingly, although these proteins are thought to play a similar role in the adhesion process, the structure of Bc28.1 from B. canis appears unrelated to the previously published structure of Bd37 from B. divergens. Site-directed mutagenesis experiments also suggest that the mechanism of the interaction with the erythrocyte membrane could be different for the two proteins. The resolution of the structure of Bc28 represents a milestone for the characterization of the parasite erythrocyte binding and its interaction with
Springer, Andrea; Fichtel, Claudia; Calvignac-Spencer, Sébastien; Leendertz, Fabian H.; Kappeler, Peter M.
Hemoparasites can cause serious morbidity in humans and animals and often involve wildlife reservoirs. Understanding patterns of hemoparasite infections in natural populations can therefore inform about emerging disease risks, especially in the light of climate change and human disruption of natural ecosystems. We investigated the effects of host age, sex, host group size and season on infection patterns of Plasmodium sp., Babesia sp. and filarial nematodes in a population of wild Malagasy primates, Verreaux's sifakas (Propithecus verreauxi), as well as the effects of these infections on hematological variables. We tested 45 blood samples from 36 individuals and identified two species of Plasmodium, one species of Babesia and two species of filarial nematodes. Plasmodium spp. and Babesia sp. infections showed opposite patterns of age-dependency, with babesiosis being prevalent among young animals, while older animals were infected with Plasmodium sp. In addition, Babesia sp. infection was a statistically significant negative predictor of Plasmodium sp. infection. These results suggest that Plasmodium and Babesia parasites may interact within the host, either through cross-immunity or via resource competition, so that Plasmodium infections can only establish after babesiosis has resolved. We found no effects of host sex, host group size and season on hemoparasite infections. Infections showed high prevalences and did not influence hematological variables. This preliminary evidence supports the impression that the hosts and parasites considered in this study appear to be well-adapted to each other, resulting in persistent infections with low pathogenic and probably low zoonotic potential. Our results illustrate the crucial role of biodiversity in host-parasite relationships, specifically how within-host pathogen diversity may regulate the abundance of parasites. PMID:26767166
Yang, Yin-Shan; Murciano, Brice; Moubri, Karina; Cibrelus, Prisca; Schetters, Theo; Gorenflot, André; Delbecq, Stéphane; Roumestand, Christian
Babesiosis (formerly known as piroplasmosis) is a tick-borne disease caused by the intraerythrocytic development of protozoa parasites from the genus Babesia. Like Plasmodium falciparum, the agent of malaria, or Toxoplasma gondii, responsible for human toxoplasmosis, Babesia belongs to the Apicomplexa family. Babesia canis is the agent of the canine babesiosis in Europe. Clinical manifestations of this disease range from mild to severe and possibly lead to death by multiple organ failure. The identification and characterization of parasite surface proteins represent major goals, both for the understanding of the Apicomplexa invasion process and for the vaccine potential of such antigens. Indeed, we have already shown that Bd37, the major antigenic adhesion protein from Babesia divergens, the agent of bovine babesiosis, was able to induce complete protection against various parasite strains. The major merozoite surface antigens of Babesia canis have been described as a 28-kDa membrane protein family, anchored at the surface of the merozoite. Here, we demonstrate that Bc28.1, a major member of this multigenic family, is expressed at high levels at the surface of the merozoite. This protein is also found in the parasite in vitro culture supernatants, which are the basis of effective vaccines against canine babesiosis. We defined the erythrocyte binding function of Bc28.1 and determined its high resolution solution structure using NMR spectroscopy. Surprisingly, although these proteins are thought to play a similar role in the adhesion process, the structure of Bc28.1 from B. canis appears unrelated to the previously published structure of Bd37 from B. divergens. Site-directed mutagenesis experiments also suggest that the mechanism of the interaction with the erythrocyte membrane could be different for the two proteins. The resolution of the structure of Bc28 represents a milestone for the characterization of the parasite erythrocyte binding and its interaction with
Understanding immune responses of white-tailed deer (WTD) to infection with Mycobacterium bovis provides insight into mechanisms of pathogen control and may provide clues to development of effective vaccine strategies. WTD were vaccinated with either BCG strain Pasteur or BCG Danish. Both vaccinates...
Monitoring serum antibody production kinetics to multiple mycobacterial antigens can be useful as a diagnostic tool for the detection of Mycobacterium bovis infection as well as for the characterization of disease progression and efficacy of intervention strategies in several species. Humoral immun...
Monitoring serum antibody production kinetics to multiple mycobacterial antigens can be useful as a diagnostic tool for the detection of Mycobacterium bovis infection as well as for the characterization of disease progression and efficacy of intervention strategies in several species. In the presen...
Nde, Chantal W; Toghrol, Freshteh; Jang, Hyeung-Jin; Bentley, William E
Tuberculosis is a leading cause of death worldwide and infects thousands of Americans annually. Mycobacterium bovis causes tuberculosis in humans and several animal species. Peracetic acid is an approved tuberculocide in hospital and domestic environments. This study presents for the first time the transcriptomic changes in M. bovis BCG after treatment with 0.1 mM peracetic acid for 10 and 20 min. This study also presents for the first time a comparison among the transcriptomic responses of M. bovis BCG to three oxidative disinfectants: peracetic acid, sodium hypochlorite, and hydrogen peroxide after 10 min of treatment. Results indicate that arginine biosynthesis, virulence, and oxidative stress response genes were upregulated after both peracetic acid treatment times. Three DNA repair genes were downregulated after 10 and 20 min and cell wall component genes were upregulated after 20 min. The devR-devS signal transduction system was upregulated after 10 min, suggesting a role in the protection against peracetic acid treatment. Results also suggest that peracetic acid and sodium hypochlorite both induce the expression of the ctpF gene which is upregulated in hypoxic environments. Further, this study reveals that in M. bovis BCG, hydrogen peroxide and peracetic acid both induce the expression of katG involved in oxidative stress response and the mbtD and mbtI genes involved in iron regulation/virulence.
Liu, Chunfa; Yue, Ruichao; Yang, Yang; Cui, Yongyong; Yang, Lifeng; Zhao, Deming; Zhou, Xiangmei
Mycobacteria can trigger the AIM2 inflammasome, autophagy activation and type-I interferon release, which are both activated by cytosolic DNA. We have recently demonstrated that activation of the AIM2 inflammasome during M. bovis infection is the result of mycobacterial translocation into the cytosol. To elucidate the effects of inflammasome activation on autophagy, we investigated the role of the AIM2 inflammasome from macrophages infected with a virulent strain of M. bovis. The results showed that the M. bovis-induced AIM2 inflammasome activation decreases autophagy in immortalized and primary murine macrophages. This relied on the inflammasome sensor AIM2 which conjugates with cytosolic DNA to inhibit the STING-dependent pathway involved in selective autophagy and interferon-β release in Mycobacterium-infected macrophages. These results suggest that the AIM2 cytosolic DNA sensor may conjugate competitively with cytosolic M. bovis DNA to restrict M. bovis induced STING-TBK1-dependent autophagy activation and IFN-β secretion. PMID:27409673
Mackintosh, C G; Qureshi, T; Waldrup, K; Labes, R E; Dodds, K G; Griffin, J F
Tuberculosis (Tb) caused by Mycobacterium bovis is a worldwide threat to livestock and humans. One control strategy is to breed livestock that are more resistant to Mycobacterium bovis. In a 3-year heritability study 6 farmed red deer stags were selected from 39 on the basis of their differing responses to experimental challenge via the tonsillar sac with approximately 500 CFU of M. bovis. Two stags remained uninfected, two were moderately affected, and two developed serious spreading Tb. Seventy offspring, bred from these six stags by artificial insemination using stored semen, were similarly challenged with M. bovis. The offspring showed patterns of response to M. bovis challenge similar to those of their sires, providing evidence for a strong genetic basis to resistance to Tb, with an estimated heritability of 0.48 (standard error, 0.096; P < 0. 01). This is the first time the heritability of Tb resistance in domestic livestock has been measured. The breeding of selection lines of resistant and susceptible deer will provide an ideal model to study the mechanisms of Tb resistance in a ruminant and could provide an additional strategy for reducing the number and severity of outbreaks of Tb in farmed deer herds. Laboratory studies to identify genetic and immunological markers for resistance to Tb are under way. Preliminary studies showed no associations between NRAMP or DRB genes and resistance to Tb in deer. Patterns of immune responses seen in resistant animals suggest that both innate and acquired pathways of immunity are necessary to produce the resistant phenotype.
Abdala, Alejandro A; Garbaccio, Sergio; Zumárraga, Martín; Tarabla, Héctor D
Control eradication campaigns of bovine tuberculosis based on the «test and slaughter» approach were successful in many countries and regions; however, in some areas the infection persists and one of the main reasons is Mycobacterium bovis infection in wild life species. Argentina has applied the same approach since 1999, achieving progress in dairy cattle herds. Nonetheless, the wildlife role has never been investigated. The objective of this study was to determine if wildlife from the Santa Fe dairy area is infected with M. bovis. Wildlife species having a positive tuberculin skin test were captured in five dairy farms. Ninety five wildlife mammals were captured; M. bovis was recovered from 7 possums (Didelphys albiventris), from one fox (Lycolapex gimnocercus) and from one rat (Rattus norvegicus). None of the animals exhibited macroscopic lesions. The most frequently isolated M. bovis spoligotypes were types 34 (4 isolates) and 12 (3 isolates). Spoligotype 34 is the most frequently isolated type in Argentine cattle. The role of D. albiventris as spillover host of M. bovis is discussed in this study.
Jang, Yunho; Ryoo, Soyoon; Lee, Hyunkyoung; Kim, Narae; Lee, Hang; Park, So-Young; Song, Woong-Seog; Kim, Jong-Taek; Lee, Hee Soo; Myung Kim, Jae
We demonstrate Mycobacterium bovis infection in wild boar ( Sus scrofa ) in South Korea. During 2012-15, we attempted to isolate M. bovis from 847 wild animals, mainly Korean water deer ( Hydropotes inermis argyropus), raccoon dogs ( Nyctereutes procyonoides ), and wild boar, from 11 regions in South Korea. We isolated M. bovis from three of 118 wild boar (2.5%) captured in Gyeonggi Province, where bovine tuberculosis (bTB) outbreaks have also occurred in livestock. Spoligotypes and mycobacterial interspersed repetitive units-variable number tandem repeats types of these M. bovis isolates (SB0140 and SB1040, 4-2-3-3-7-5-5-4-4-3-4-3 and 5-2-3-3-7-5-5-4-3-10-5-2; MIRU4, MIRU16, MIRU27, MIRU31, ETR-A, ETR-B, ETR-C, QUB11b, QUB26, QUB3336, VNTR2401, and VNTR3171) have also been identified from farmed livestock such as cattle ( Bos taurus coreanae), Formosan sika deer ( Cervus nippon taiouanus), and American elk ( Cervus canadensis ) in the country. In South Korea, bTB appears to be endemic in livestock, and there are numerous opportunities for contact between wild boar and livestock due to high population densities and broad activity ranges. Our results support the hypothesis that M. bovis is transmitted between domestic and wild animals.
Dyer, Neil; Register, Karen B; Miskimins, Dale; Newell, Teresa
Mycoplasma bovis has emerged as a significant and costly infectious disease problem in bison, generally presenting as severe, caseonecrotic pneumonia. Three diagnostic cases in which M. bovis is associated with necrotic pharyngitis in bison are described in the current study. The bacterium was isolated from lesions of the pharynx or lung of 3 American bison (Bison bison), at 2 different locations in the upper Midwestern United States, with severe, necrotic pharyngeal abscesses. Chronic caseonecrotic inflammation typical of M. bovis infection in bovines was observed microscopically in the pharynxes of affected bison. A mixed population of bacteria was recovered from the pharyngeal lesions, and Trueperella pyogenes, a frequent secondary pathogen in ruminant respiratory disease, was consistently isolated from the affected animals. Distinctive histopathological features of the pharyngeal lesions favor causation by M. bovis, although a role for T. pyogenes in the clinical presentation cannot be excluded. Veterinarians and producers working with bison should be aware that M. bovis may be associated with pharyngitis in bison.
Suleman, Muhammad; Prysliak, Tracy; Windeyer, Claire; Perez-Casal, Jose
Mycoplasma bovis is a pathogen globally affecting cattle and bison herds, causing pneumonia, arthritis, mastitis, abortions, and other symptoms, leading to huge economic losses. Many studies have been done regarding the antimicrobial susceptibility of M. bovis isolated from cattle, but no such study is available for isolates recovered from bison. For the first time, in vitro susceptibilities of 40 M. bovis clinical isolates collected from bison herds in Canada are reported here. Minimal inhibitory concentration (MIC) values were determined using Sensititre® plates. The most effective MIC50 and MIC90 were for spectinomycin (1 and >64 μg/mL), tiamulin (1 and >32 μg/mL), and tulathromycin (16 and 64 μg/mL), whereas tetracyclines, fluoroquinolones, and florfenicol failed to inhibit growth of M. bovis bison isolates. Isolates were nonsusceptible to tetracyclines (100%), fluoroquinolones (97.5%), and tilmicosin (100%), whereas the highest susceptibility of bison clinical isolates was seen with spectinomycin (95%) and tulathromycin (67.5%). Two lung isolates (Mb283 and 348) were found resistant to both spectinomycin and tulathromycin. These results show a marked difference in antimicrobial susceptibility of bison isolates as compared with previously reported and laboratory reference cattle isolates, emphasizing the necessity of testing antimicrobial susceptibility of M. bovis bison isolates and to generate better therapeutic regime for improved recovery chances for infected bison herds across North America.
Register, Karen B; Thole, Luke; Rosenbush, Ricardo F; Minion, F Chris
Mycoplasma bovis is a primary agent of mastitis, pneumonia and arthritis in cattle and the bacterium most frequently isolated from the polymicrobial syndrome known as bovine respiratory disease complex. Recently, M. bovis has emerged as a significant health problem in bison, causing necrotic pharyngitis, pneumonia, dystocia and abortion. Whether isolates from cattle and bison comprise genetically distinct populations is unknown. This study describes the development of a highly discriminatory multilocus sequencing typing (MLST) method for M. bovis and its use to investigate the population structure of the bacterium. Genome sequences from six M. bovis isolates were used for selection of gene targets. Seven of 44 housekeeping genes initially evaluated were selected as targets on the basis of sequence variability and distribution within the genome. For each gene target sequence, four to seven alleles could be distinguished that collectively define 32 sequence types (STs) from a collection of 94 cattle isolates and 42 bison isolates. A phylogeny based on concatenated target gene sequences of each isolate revealed that bison isolates are genetically distinct from strains that infect cattle, suggesting recent disease outbreaks in bison may be due to the emergence of unique genetic variants. No correlation was found between ST and disease presentation or geographic origin. MLST data reported here were used to populate a newly created and publicly available, curated database to which researchers can contribute. The MLST scheme and database provide novel tools for exploring the population structure of M. bovis and tracking the evolution and spread of strains.
Kelly, Pamela; Jahns, Hanne; Power, Eugene; Bainbridge, John; Kenny, Kevin; Corpa, Juan M; Cassidy, Joseph P; Callanan, John J
Avian tuberculosis rarely affects ratites compared to other bird species and is typically caused by Mycobacterium avium species. This study describes the pathological and microbiological findings in three adult ostriches with mycobacteriosis, in one of which Mycobacterium bovis was isolated from the lesions. Post mortem examinations on ostriches from two different zoological collections in Ireland revealed multifocal caseous granulomas affecting the spleen and liver in all cases, with additional involvement of intestines in two cases. In one case, granulomas were present within the pharynx, at the thoracic inlet and multifocally on the pleural surface. Acid-fast bacilli were observed in all lesions. Mycobacterium sp. of the M. avium complex was isolated from the intestinal lesions in the two cases with intestinal involvement, and M. bovis sp. oligotype SB0140 was cultured from the liver of the third ostrich. This represents the first reported case of M. bovis infection in an ostrich. Avian tuberculosis due to M. bovis is rare and to date has been reported in only parrots and experimentally inoculated birds. Mycobacterium bovis needs to be considered as a possible cause of tuberculosis in ostriches because the lesions are similar to those observed with M. avium complex infection.
Scott, Colleen; Cavanaugh, Joseph S; Silk, Benjamin J; Ershova, Julia; Mazurek, Gerald H; LoBue, Philip A; Moonan, Patrick K
Current US guidelines recommend longer treatment for tuberculosis (TB) caused by pyrazinamide-resistant organisms (e.g., Mycobacterium bovis) than for M. tuberculosis TB. We compared treatment response times for patients with M. bovis TB and M. tuberculosis TB reported in the United States during 2006-2013. We included culture-positive, pulmonary TB patients with genotyping results who received standard 4-drug treatment at the time of diagnosis. Time to sputum-culture conversion was defined as time between treatment start date and date of first consistently culture-negative sputum. We analyzed 297 case-patients with M. bovis TB and 30,848 case-patients with M. tuberculosis TB. After 2 months of treatment, 71% of M. bovis and 65% of M. tuberculosis TB patients showed conversion of sputum cultures to negative. Likelihood of culture conversion was higher for M. bovis than for M. tuberculosis, even after controlling for treatment administration type, sex, and a composite indicator of bacillary burden.
Mon, María Laura; Moyano, Roberto Damián; Viale, Mariana Noelia; Colombatti Olivieri, María Alejandra; Gamietea, Ignacio José; Montenegro, Valeria Noely; Alonso, Bernardo; Santangelo, María de la Paz; Singh, Mahavir; Duran, Rosario; Romano, María Isabel
The Delayed type hypersensitivity skin test (DTH) and interferon-gamma assay are used for the diagnosis of bovine tuberculosis (TBB). The specificity of these diagnoses, however, is compromised because both are based on the response against purified protein derivative of Mycobacterium bovis (PPD-B). In this study, we assessed the potential of two cocktails containing M. bovis recombinant proteins: cocktail 1 (C1): ESAT-6, CFP-10 and MPB83 and cocktail 2 (C2): ESAT-6, CFP-10, MPB83, HspX, TB10.3, and MPB70. C1, C2, and PPD-B showed similar response by DTH in M. bovis-sensitized guinea pigs. Importantly, C1 induced a lower response than PPD-B in M. avium-sensitized guinea pigs. In cattle, C1 displayed better performance than PPD-B and C2; indeed, C1 showed the least detection of animals either vaccinated or Map-infected. To optimize the composition of the cocktails, we obtained protein fractions from PPD-B and tested their immunogenicity in experimentally M. bovis-infected cattle. In one highly reactive fraction, seven proteins were identified. The inclusion of FixB in C1 enhanced the recognition of M. bovis-infected cattle without compromising specificity. Our data provide a promising basis for the future development of a cocktail for TBB detection without interference by the presence of sensitized or infected animals with other mycobacteria. PMID:25110654
Michel, A L; Coetzee, M L; Keet, D F; Maré, L; Warren, R; Cooper, D; Bengis, R G; Kremer, K; van Helden, P
Bovine tuberculosis is endemic in African buffalo and a number of other wildlife species in the Kruger National Park (KNP) and Hluhluwe-iMfolozi Park (HiP) in South Africa. It was thought that the infection had been introduced into the KNP ecosystem through direct contact between cattle and buffalo, a hypothesis which was confirmed in this study by IS6110 and PGRS restriction fragment length polymorphism (RFLP) typing. The molecular characterisation of 189 Mycobacterium bovis isolates from nine wildlife species in the HiP, including three smaller associated parks, and the Kruger National Park with adjacent areas showed that the respective epidemics were each caused by an infiltration of a single M. bovis genotype. The two M. bovis strains had different genetic profiles, as demonstrated by hybridisation with the IS6110 and PGRS RFLP probes, as well as with regard to evidence of evolutionary changes to the IS profile. While the M. bovis type in HiP was transmitted between buffaloes and to at least baboon, bushpig and lion without obvious genetic changes in the RFLP patterns, in the KNP a dominant strain was represented in 73% of the M. bovis isolates, whilst the remaining 27% were variants of this strain. No species-specific variants were observed, except for one IS6110 type which was found only in a group of five epidemiologically related greater kudu. This finding was attributed to species-specific behaviour patterns rather than an advanced host-pathogen interaction.
Behrendt, Jan Hillern; Ruiz, Antonio; Zahner, Horst; Taubert, Anja; Hermosilla, Carlos
Eimeria bovis infections are under immunological control and recent studies have emphasized the role of early PMN-mediated innate immune responses in infected calves. Neutrophil extracellular traps (NETs) have recently been demonstrated to act as a killing mechanism of PMN against several pathogens. In the present study, the interactions of bovine PMN with sporozoites of E. bovis were investigated in this respect in vitro. For demonstration and quantification of NET formation, extracellular DNA was stained by Sytox Orange. Fluorescence images after Sytox Orange staining as well as scanning electron microscopy (SEM) showed NET formation to occur upon contact with E. bovis sporozoites. Exposure of PMN to viable sporozoites induced stronger NET formation than to dead or homogenized parasites. NET formation was abolished by treatment with DNase and could be reduced by diphenylene iodonium, which is described as a potent inhibitor of NADPH oxidase. After sporozoite and PMN co-culture, extracellular fibres were found attached to sporozoites and seemed to trap them, strongly suggesting that NETs immobilize E. bovis sporozoites and thereby prevent them from infecting host cells. Thus, transfer of sporozoites, previously being confronted with PMN, to adequate host cells resulted in clearly reduced infection rates when compared to PMN-free controls. NET formation by PMN may therefore represent an effector mechanism in early innate immune reactions against E. bovis. This is the first report indicating Eimeria-induced NET formation.
Bangoura, Berit; Mundt, Hans-Christian; Schmäschke, Ronald; Westphal, Bernhard; Daugschies, Arwid
The present study was designed to investigate the prevalence of the pathogenic coccidia species Eimeria bovis and Eimeria zuernii in shed-reared animals in German dairy and fattening facilities. Samples were obtained from 65 cattle farms distributed randomly across all the regions of Germany regardless of the occurrence of clinical problems. The samples were obtained rectally. Faecal consistency and the total number of oocysts per gram of faeces (OPG) were determined, along with the OPG values for E. bovis and E. zuernii. A questionnaire was completed for each farm to record information about herd size and management, along with individual animal data. Eimeria oocysts were detected in 62 of these farms, which give a prevalence of 95.4%. The farm prevalence of the pathogenic species was 76.9% for E. bovis and 83.1% for E. zuernii. The number of oocysts excreted could not be correlated significantly with farm type or farm management but depended on the floor type, the age of the calves and the time after rehousing. Furthermore, there was a positive correlation between OPG and the observation of diarrhoea. E. zuernii had a greater influence on the occurrence of diarrhoea than E. bovis. This study confirms that herd management frequently does not meet the requirements of effective coccidia control despite the fact that the pathogenic coccidia E. bovis and E. zuernii are ubiquitous in German cattle populations.
Cavanaugh, Joseph S.; Silk, Benjamin J.; Ershova, Julia; Mazurek, Gerald H.; LoBue, Philip A.; Moonan, Patrick K.
Current US guidelines recommend longer treatment for tuberculosis (TB) caused by pyrazinamide-resistant organisms (e.g., Mycobacterium bovis) than for M. tuberculosis TB. We compared treatment response times for patients with M. bovis TB and M. tuberculosis TB reported in the United States during 2006–2013. We included culture-positive, pulmonary TB patients with genotyping results who received standard 4-drug treatment at the time of diagnosis. Time to sputum-culture conversion was defined as time between treatment start date and date of first consistently culture-negative sputum. We analyzed 297 case-patients with M. bovis TB and 30,848 case-patients with M. tuberculosis TB. After 2 months of treatment, 71% of M. bovis and 65% of M. tuberculosis TB patients showed conversion of sputum cultures to negative. Likelihood of culture conversion was higher for M. bovis than for M. tuberculosis, even after controlling for treatment administration type, sex, and a composite indicator of bacillary burden. PMID:28221125
Johansen, Tone Bjordal; Muma, John Bwalya; Munyeme, Musso; Mbulo, Grace; Muwonge, Adrian; Djønne, Berit
Tuberculosis remains a major public health problem in Zambia. While human to human transmission of Mycobacterium tuberculosis is of major importance in driving the tuberculosis epidemic, the impact of Mycobacterium bovis transmission from infected cattle is largely unknown. This cross-sectional study aimed at molecular characterization of M. bovis in humans and cattle. A total of 100 human sputum samples and 67 bovine tissues were collected and analyzed for the presence of mycobacteria. Of 65 human samples that harbored acid fast bacteria (AFB), 55 isolates were obtained of which 34 were identified as M. tuberculosis and 2 as M. bovis. AFB-positive bovine samples (n = 67) yielded 47 mycobacterial isolates among which 25 were identified as M. bovis and no M. tuberculosis was found. Among the M. bovis isolates, spoligotyping revealed a high homogeneity in genotypes circulating in Namwala district. Human and cattle isolates shared identical MIRU-VNTR genotypes, suggesting that transmission between the two hosts may occur. Therefore, this study has documented zoonotic TB in human patients in Namwala district of Zambia. However, further molecular epidemiological studies in the study area are recommended. PMID:24847441
Malama, Sydney; Johansen, Tone Bjordal; Muma, John Bwalya; Munyeme, Musso; Mbulo, Grace; Muwonge, Adrian; Djønne, Berit; Godfroid, Jacques
Tuberculosis remains a major public health problem in Zambia. While human to human transmission of Mycobacterium tuberculosis is of major importance in driving the tuberculosis epidemic, the impact of Mycobacterium bovis transmission from infected cattle is largely unknown. This cross-sectional study aimed at molecular characterization of M. bovis in humans and cattle. A total of 100 human sputum samples and 67 bovine tissues were collected and analyzed for the presence of mycobacteria. Of 65 human samples that harbored acid fast bacteria (AFB), 55 isolates were obtained of which 34 were identified as M. tuberculosis and 2 as M. bovis. AFB-positive bovine samples (n = 67) yielded 47 mycobacterial isolates among which 25 were identified as M. bovis and no M. tuberculosis was found. Among the M. bovis isolates, spoligotyping revealed a high homogeneity in genotypes circulating in Namwala district. Human and cattle isolates shared identical MIRU-VNTR genotypes, suggesting that transmission between the two hosts may occur. Therefore, this study has documented zoonotic TB in human patients in Namwala district of Zambia. However, further molecular epidemiological studies in the study area are recommended.
Kieser, T; Moss, M T; Dale, J W; Hopwood, D A
The ability of "Streptomyces lividans" to use the expression signals of genes from Mycobacterium bovis BCG was tested in vivo by using gene fusions. Random DNA fragments from M. bovis BCG were inserted into promoter-probe plasmids in Escherichia coli and in "S. lividans." Comparison with promoter activity detected with random DNA fragments from the respective hosts suggested that "S. lividans" efficiently utilizes a high proportion of mycobacterial promoters, whereas a smaller fraction are expressed, and expressed more weakly, in E. coli. M. bovis BCG DNA fragments were also inserted into the specially constructed translational fusion vector (pIJ688) in "S. lividans." pIJ688 contains the kanamycin phosphotransferase gene (neo) from transposon Tn5, truncated at its amino terminus, as the indicator. The results suggested that "S. lividans" uses M. bovis BCG translational signals almost as efficiently as its own signals. Moreover, several hybrid proteins with an M. bovis BCG-derived amino terminus seemed to be reasonably stable in "S. lividans." These experiments indicate that "S. lividans" may be a suitable host for the expression of Mycobacterium leprae and Mycobacterium tuberculosis genes from their own signals. This is a precondition for the expression of entire biosynthetic pathways, which could be valuable in the production of diagnostic and therapeutic agents. The vectors may also have wider applications for the analysis of gene expression in Streptomyces.
Nalubamba, King Shimumbo; Mudenda, Ntombi Basimbi; Namwila, Mwaka Mwangala; Mulenga, Chilufya Susan; Bwalya, Eugene Chisela; M'kandawire, Ethel; Saasa, Ngonda; Hankanga, Careen; Oparaocha, Elizabeth; Simuunza, Martin
A retrospective and prospective analysis of clinical records of dogs diagnosed with Babesia infections was carried out for the years 2000 to 2013 from practices in Lusaka, Zambia. Records of 363 dogs with confirmed Babesia infections were analysed using demographic factors including sex, breed, age, and clinical signs in relation to haematological findings and Babesia species. The clinical and laboratory findings observed are described as well as Babesia species identification. The study included 18 breeds and the highest proportion were mongrels (32.2%), males representing 64.5% of the population. The most common presenting problems were anorexia (65.3%) and lethargy/weakness (65.3%). The most common clinical signs were fever (87.3%), pallor (52.3%), lymphadenopathy (47.4%), and presence of ticks (44.9%). Anaemia (96.4%) and nucleated erythrocytes (42.2%) were the most common laboratory findings. A mixed infection of Babesia rossi and Babesia gibsoni was present in 59.7% of dogs, whilst 8% and 32.2% had B. rossi and B. gibsoni as a single infection, respectively. Case management mainly involved therapy with tetracyclines and imidocarb and was usually accompanied by clinical improvement. This study highlights, for the first time, the presence of B. gibsoni in natural dog populations in Zambia, where previously only B. rossi was reported. PMID:26682062
Mycoplasma bovis has recently emerged as a significant and costly infectious disease problem in bison. This report demonstrates that ELISAs for detection of M. bovis-specific antibody in cattle are not optimal for identification of seropositive bison. An ELISA optimized for use with bison sera is ...
Numerous species of wild mammals are susceptible to Mycobacterium bovis, a cause of bovine tuberculosis (TB). Eurasian badgers, white-tailed deer, brushtail possums, and wild boar are implicated in the maintenance of wildlife reservoirs of M. bovis infection in different countries, fueling bovine TB...
Wildlife reservoirs of Mycobacterium bovis represent serious obstacles to the eradication of tuberculosis from livestock, particularly cattle. In Michigan, USA tuberculous white-tailed deer transmit M. bovis to other deer and cattle. One approach in dealing with this wildlife reservoir is to vaccina...
Milian-Suazo, Feliciano; Harris, Beth; Arriaga Díaz, Camila; Romero Torres, Cecilia; Stuber, Tod; Alvarez Ojeda, Genoveva; Morales Loredo, Alberto; Perez Soria, Martina; Payeur, Janet B
Tuberculosis (TB) represents a barrier for free trade of livestock between Mexico and the United States of America (US). In spite of efforts from Mexico to export TB-free animals, some of those found with TB lesions in slaughterhouses in the US are traced back to that country. Therefore, the purpose of this study was to determine, through molecular epidemiology, the most probable source of infection for cattle found with TB lesions in the US. Ninety M. bovis isolates, 50 from Mexico obtained from cattle in 8 different states, and 40 from the US from cattle, deer, elk and feral pigs from 7 different states were included in the study. All samples were analyzed in both laboratories, Mexico and the US, following the same protocol for molecular analysis by spoligotyping. Twenty-seven clusters, ranging from 1 to 18 genetically similar strains were found. Some clustering by country was observed, strains from cattle and deer in Michigan in the US fell into the same cluster, suggesting transmission between species. These results, combined with epidemiological information suggest that despite of the possibility that some animals with lesions in the US come from Mexico as false negatives, the US has its own source of infection, must probably in dairy cattle and wildlife. Genetic diversity of isolates from Mexico was larger than that in the US, which could be a consequence of the endemic status of the disease and the indiscriminate movement of animals between regions.
Brooks, B W; Barnum, D A
Twenty bovine udder quarters colonized with Corynebacterium bovis SR6 and 20 uncolonized quarters were challenged by inoculation of Staphylococcus aureus Newbould 305 (ATCC 29740) into the teat cistern. The percentage of infection in quarters colonized with C. bovis (50%) was significantly lower than that in controls (100%). By similar challenge no significant difference was observed between the percentage of infection with Streptococcus agalactiae ATCC 27956 in 33 quarters colonized with C. bovis (70%) compared to 33 controls (87.9%). A total of 37 quarters colonized with C. bovis and 37 control quarters were challenged with Staph. aureus Newbould 305 (ATCC 29740) and Maxi (ATCC 27543) and Strep. agalactiae (ATCC 27956) by exposure of the teat orifice. The percentage of teat ducts colonized with C. bovis which became infected with either pathogen was not different from that for controls. PMID:6372969
Parra, A; García, A; Inglis, N F; Tato, A; Alonso, J M; Hermoso de Mendoza, M; Hermoso de Mendoza, J; Larrasa, J
Recreational hunting of indigenous wild artiodactyls has been one of the most lucrative and rapidly growing industries in Western Spain over the last five years. In the absence of careful ecological management, one consequence of the commercial exploitation of this natural resource has been the appearance of outbreaks of infectious disease; most notably bovine tuberculosis. From the outset of the study in 1997, we have observed a steady increase in prevalence of Mycobacterium bovis (M. bovis) in both species reaching 1.74 (+/-0.17) in deer in 2002 and 2.32 (+/-0.24) in wild boar. The latter species seems to be most severely affected with pulmonary lesions appearing more chronic than those observed in deer. In this study, we describe the epidemiology of M. bovis in European wild boar (Sus scrofa) and Iberian red deer (Cervus elaphus hispanicus) in Extremadura (W. Spain); a region where there are large areas of natural habitat for these species.
Balseiro, Ana; Rodríguez, Oscar; González-Quirós, Pablo; Merediz, Isabel; Sevilla, Iker A; Davé, Dipesh; Dalley, Deanna J; Lesellier, Sandrine; Chambers, Mark A; Bezos, Javier; Muñoz, Marta; Delahay, Richard J; Gortázar, Christian; Prieto, José M
The prevalence, distribution and pathology related to infection with Mycobacterium bovis and other mycobacteria were determined in trapped (n=36) and road-killed (n=121) badgers in Spain from 2006 to 2010. The prevalence of M. bovis based on bacteriological culture from road-killed badgers was 8/121 (6.6%) and from trapped badgers was 0/36 (0%). Tuberculosis/M. bovis infection was evident in 15/121 (12.4%) road-killed badgers when bacteriology and histopathology were combined. Mycobacterium avium complex was isolated by culture from the tracheal aspirate of 1/36 (2.8%) trapped badgers and from tissue pools from 8/121 (6.6%) road-killed badgers.
Aljada, Ibrahim S.; Crane, John K.; Corriere, Nancy; Wagle, Datta G.; Amsterdam, Daniel
We report a case of Mycobacterium bovis BCG vertebral osteomyelitis in a 79-year-old man 2.5 years after intravesical BCG therapy for bladder cancer. The recovered isolate resembled M. tuberculosis biochemically, but resistance to pyrazinamide (PZA) rendered that diagnosis suspect. High-pressure liquid chromatographic studies confirmed the diagnosis of M. bovis BCG infection. The patient was originally started on a four-drug antituberculous regimen of isoniazid, rifampin, ethambutol, and PZA. When susceptibility studies were reported, the regimen was changed to isoniazid and rifampin for 12 months. Subsequently, the patient was transferred to a skilled nursing facility for 3 months, where he underwent intensive physical therapy. Although extravesical adverse reactions are rare, clinicians and clinical microbiologists need to be aware of the possibility of disseminated infection by M. bovis BCG in the appropriate setting of clinical history, physical examination, and laboratory investigation. PMID:10325395
Kakuda, Tsutomu; Sarataphan, Nopporn; Tanaka, Tetsuya; Takai, Shinji
The complete nucleotide sequence of a plasmid, pMBO-1, from Moraxella bovis strain Epp63 was determined. We identified 30 open reading frames (ORFs) encoded by the 44,215bp molecule. Two large ORFs, flpA and flpB, encoding proteins with similarity to Bordetella pertussis filamentous haemagglutinin (FHA), were identified on the same plasmid. The gene for a specific accessory protein (Fap), which may play a role in the secretion of Flp protein, was also identified. Reverse transcriptase PCR analysis of total RNA isolated from M. bovis Epp63 indicated that the flpA, flpB, and fap genes are all transcribed. Southern blot analysis indicated that the flp and fap genes are present in other clinical isolates of geographically diverse M. bovis.
Hermosilla, Carlos; Schröpfer, Elmar; Stowasser, Michael; Eckstein-Ludwig, Ursula; Behrendt, Jan Hillern; Zahner, Horst
The first merogony of Eimeria bovis takes place in lymphatic endothelial cells of the ileum, resulting in the formation of macromeronts up to 250 microm. In this study, we investigated the host cell cytoskeleton (actin filaments, microtubules, spectrin, vimentin intermediate filaments) associated with parasitic development in vitro by confocal laser scanning microscopy (CLSM) using primary bovine umbilical vein endothelial cells (BUVEC) and bovine spleen lymphatic endothelial cells (BSLEC) as host cells. No prominent changes in the host cell cytoskeleton were detected 1-3 days after E. bovis sporozoite invasion. With ongoing meront maturation a significant increase in microtubules and actin filaments close to the parasitophorous vacuole (PV) was found. Mature macromeronts within the PV were completely enclosed by these cytoskeletal elements. Our findings suggest, that in order to guarantee the survival of the host cell on the enlargement of macromeronts, E. bovis needs not only to augment but also to rearrange its cytoskeletal system.
Behrendt, J H; Clauss, W; Zahner, H; Hermosilla, C
In vitro Eimeria bovis sporozoites invade a wide range of cell types, and in the case of bovine cells, they may develop to first-generation schizonts. Often, however, they subsequently leave their host cell to invade a new one, which seems contrary to the classical way of infecting a cell by forming a parasitophorous vacuole. Using a standard, "cell wound assay," we show that E. bovis can invade bovine endothelial cells by breaching the plasma membrane and may again leave the surviving cell. Eimeria bovis sporozoites also infected VERO and HT29 cells but obviously without damaging the plasma membrane. The same held true when bovine endothelial cells were exposed to tachyzoites of Toxoplasma gondii and Neospora caninum. According to a literature report dealing with Plasmodium yoelii sporozoites, breaching the membrane of certain host cells may be a common phenomenon for coccidian sporozoites but may not be for merozoites.
Gross, W M; Hawkins, J E
In the context of a busy reference laboratory, radiometric selective inhibition tests were evaluated for rapid differentiation of Mycobacterium tuberculosis and Mycobacterium bovis and of the M. tuberculosis complex from other mycobacteria. p-Nitro-alpha-acetylamino-beta-hydroxypropiophenone at 5 micrograms and hydroxylamine hydrochloride at 62.5 and 125 micrograms per ml of 7H12 medium were used to separate the M. tuberculosis complex from other mycobacteria (MOTT bacilli). Since it is important epidemiologically to distinguish M. tuberculosis from M. bovis, susceptibility to 1 microgram of thiophene-2-carboxylic acid per ml was also determined radiometrically. By using these three agents as selective inhibitors, M. tuberculosis, M. bovis, and MOTT bacilli were differentiated with a high degree of specificity by a BACTEC radiometric procedure. Results of tests performed on clinical isolates submitted on solid medium to our reference laboratory were available within 5 days. PMID:3921561
Hamid, Penny H; Hirzmann, Joerg; Kerner, Katharina; Gimpl, Gerald; Lochnit, Guenter; Hermosilla, Carlos R; Taubert, Anja
During first merogony Eimeria bovis forms large macromeronts in endothelial host cells containing >120 000 merozoites I. During multiplication, large amounts of cholesterol are indispensable for the enormous offspring membrane production. Cholesterol auxotrophy was proven for other apicomplexan parasites. Consequently they scavenge cholesterol from their host cell apparently in a parasite-specific manner. We here analyzed the influence of E. bovis infection on endothelial host cell cholesterol metabolism and found considerable differences to other coccidian parasites. Overall, free cholesterol significantly accumulated in E. bovis infected host cells. Furthermore, a striking increase of lipid droplet formation was observed within immature macromeronts. Artificial host cell lipid droplet enrichment significantly improved E. bovis merozoite I production confirming the key role of lipid droplet contents for optimal parasite proliferation. The transcription of several genes being involved in both, cholesterol de novo biosynthesis and low density lipoprotein-(LDL) mediated uptake, was significantly up-regulated at a time in infected cells suggesting a simultaneous exploitation of these two cholesterol acquisition pathways. E. bovis scavenges LDL-derived cholesterol apparently through significantly increased levels of surface LDL receptor abundance and LDL binding to infected cells. Consequently, LDL supplementation significantly improved parasite replication. The up-regulation of the oxidized LDL receptor 1 furthermore identified this scavenger receptor as a key molecule in parasite-triggered LDL uptake. Moreover, cellular cholesterol processing was altered in infected cells as indicated by up-regulation of cholesterol-25-hydroxylase and sterol O-acyltransferase. Overall, these results show that E. bovis considerably exploits the host cell cholesterol metabolism to guarantee its massive intracellular growth and replication.
Esquivel-Solís, Hugo; Vallecillo, Antonio J.; Benítez-Guzmán, Alejandro; Adams, L. Garry; López-Vidal, Yolanda; Gutiérrez-Pabello, José A.
To identify the resistance phenotype against Mycobacterium bovis in cattle, we used a bactericidal assay that has been considered a marker of this trait. Three of 24 cows (12.5%) were phenotyped as resistant and 21 as susceptible. Resistance of bovine macrophages (MΦ) to BCG challenge was evaluated for its association with SLC11A1 GT microsatellite polymorphisms within 3′UTR region. Twenty-three cows (95.8%) had a GT13 genotype, reported as resistant, consequently the SLC11A1polymorphism was not in agreement with our bactericidal assay results. MΦ of cows with resistant or susceptible phenotype were challenged in vitro with virulent M. bovis field strain or BCG, and nitric oxide production, bacterial killing and apoptosis induction were measured in resting and LPS-primed states. M. bovis field strain induced more apoptosis than BCG, although the difference was not significant. Resistant MΦ controlled better the replication of M. bovis (P<0.01), produced more nitric oxide (P<0.05) and were slightly more prone to undergo apoptosis than susceptible cells. LPS pretreatment of MΦ enhanced all the functional parameters analyzed. Inhibition of nitric oxide production with nG-monomethyl-L-arginine monoacetate enhanced replication of M. bovis but did not modify apoptosis rates in both resistant and susceptible MΦ. We conclude that nitric oxide production not apoptosis is a major determinant of macrophage resistance to M. bovis infection in cattle and that the influence of SLC11A1 gene 3′UTR polymorphism is not associated with this event. PMID:23691050
Corredoira, J; Alonso, M P; Coira, A; Casariego, E; Arias, C; Alonso, D; Pita, J; Rodriguez, A; López, M J; Varela, J
The purpose of this study was to evaluate the characteristics of infective endocarditis (IE) caused by S. bovis and compare them to those caused by streptococci of the viridans group (SVG). A prospective study was undertaken considering 55 consecutive cases of IE due to S. bovis and 41 to SVG over 18 years. The study was divided into two periods (1988-1996 and 1997-2005). S. bovis caused 24% of the IE in our centre and constituted the main aetiology for this disease, showing an increase of 358% during the second period studied. Biotype I was responsible for 94.5% of cases and there was a high degree of association with colon tumours (53%). Over the period of the study, 107 patients admitted to our hospital had bacteraemia caused by S. bovis and 310 patients had bacteraemia caused by SVG. In the first group, 55 (51%) were endocarditis cases, but only 41 (13%) of the patients with SVG bacteraemia had endocarditis (p < 0.0001). The distinguishing features of endocarditis caused by S. bovis in comparison with those caused by SGV were: a greater increase in cases during the 2nd period studied (from 12 to 43 vs. from 19 to 22, p < 0.01), a higher percentage of males (93% vs. 71%, p < 0.004), patients significantly older (median age 66 vs. 58.5, p < 0.004), less predisposing cardiopathy (42% vs. 76%, p < 0.0009), more bivalvular involvement (42% vs. 22%, p < 0.04), more spondylitis (9% vs. 0%, p < 0.04), a higher association with colonic tumours (53% vs. 5%, p < 0.0001), and a higher percentage of antibiotic resistance: erythromycin 66% vs. 19%, p < 0.0001; clindamycin 67% vs. 11%, p < 0.0001; cotrimoxazole 77% vs. 30.5%, p < 0.0001, respectively. IE due to S. bovis is an emergent disease in our environment, presenting different characteristics to those produced by SVG.
Al Izzi, Salah; Martin, Donald S; Chan, Roxanne Y Y; Leutenegger, Christian M
A 12-month-old male neutered mixed breed dog was presented with a history of diarrhea, lethargy, emaciation, polydypsia, and sniffling. Physical examination findings included pale mucous membranes, increased heart and respiratory rates, and normal rectal temperature (38°C). Hematologic abnormalities included anemia and thrombocytopenia. Biochemical abnormalities included hypoalbuminemia, hyperbilirubinemia, and elevated ALP and ALT activities. A SNAP 4Dx test result was positive for Ehrlichia canis. Babesia canis vogeli organisms were found in the peripheral blood films, while morulae of E canis were not seen. Real-time polymerase chain reaction testing confirmed the presence of both B c vogeli and E canis organisms, and also was positive for Anaplasma platys infection. The dog recovered following treatment with doxycycline and imidocarb dipropionate, with normal hematology and biochemical profiles.
El Imam, Ahmed H; Hassan, Shawgi M; Gameel, Ahmed A; El Hussein, Abdelrahim M; Taha, Khalid M; Oosthuizen, Marinda C
The epidemiological aspects of sheep piroplasmosis in Sudan are poorly studied, and further investigations using sensitive and precise techniques are required. In this study, the Reverse Line Blot (RLB) hybridization assay was used to detect and simultaneously differentiate between Theileria and Babesia species. DNA was extracted from blood collected on filter paper (n=219) from apparently healthy sheep from six different geographical localities in Sudan. Results indicated that Theileria ovis (88.6%), T. separata (20.1%), T. lestoquardi (16.4%) and T. annulata (16.4%) DNA could be detected in the blood samples. Single and mixed Theileria infections were detected in 74 (33.8%) and 124 (56.6%) respectively and T. ovis being the most prevalent species in the country. T. ovis and T. separata were reported for the first time in sheep in Sudan.
Jefferies, R; Ryan, U M; Jardine, J; Broughton, D K; Robertson, I D; Irwin, P J
This study reports on the epidemiology of Babesia gibsoni in American Pit Bull Terriers living in a region of western Victoria in southern Australia. Both American Pit Bull Terriers (n = 100) and other dog breeds (n = 51) were screened for B gibsoni using immunofluorescent antibody testing (IFAT) and/or polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). A questionnaire was also completed by each dog owner, ascertaining the husbandry and habits of the dogs sampled. Fourteen dogs were positive for B gibsoni using IFAT and/or PCR-RFLP and all were American Pit Bull Terriers. Dogs that were male and/or had been bitten by or were biters of other American Pit Bull Terriers were more likely to be B gibsoni positive, thus suggesting that blood-to-blood transmission contributes to the spread of this disease between dogs.
Krause, Peter J.; Spielman, Andrew; Goethert, Heidi; Gern, Lise; Brand, Brigit; Telford, Sam R.
We evaluated human risk for infection with Babesia microti at a site in eastern Switzerland where several B. microti–infected nymphal Ixodes ricinus ticks had been found. DNA from pooled nymphal ticks amplified by polymerase chain reaction was highly homologous to published B. microti sequences. More ticks carried babesial infection in the lower portion of the rectangular 0.7-ha grid than in the upper (11% vs. 0.8%). In addition, we measured seroprevalence of immunoglobulin (Ig) G antibodies against B. microti antigen in nearby residents. Serum from 1.5% of the 396 human residents of the region reacted to B. microti antigen (>1:64), as determined by indirect immunofluorescence assay (IgG). These observations constitute the first report demonstrating B. microti in a human-biting vector, associated with evidence of human exposure to this agent in a European site. PMID:12095442
Foppa, Ivo M; Krause, Peter J; Spielman, Andrew; Goethert, Heidi; Gern, Lise; Brand, Brigit; Telford, Sam R
We evaluated human risk for infection with Babesia microti at a site in eastern Switzerland where several B. microti-infected nymphal Ixodes ricinus ticks had been found. DNA from pooled nymphal ticks amplified by polymerase chain reaction was highly homologous to published B. microti sequences. More ticks carried babesial infection in the lower portion of the rectangular 0.7-ha grid than in the upper (11% vs. 0.8%). In addition, we measured seroprevalence of immunoglobulin (Ig) G antibodies against B. microti antigen in nearby residents. Serum from 1.5% of the 396 human residents of the region reacted to B. microti antigen (>1:64), as determined by indirect immunofluorescence assay (IgG). These observations constitute the first report demonstrating B. microti in a human-biting vector, associated with evidence of human exposure to this agent in a European site.
Bautista-Garfias, C R; Gómez, M B; Aguilar, B R; Ixta, O; Martínez, F; Mosqueda, J
In this study, we report that administration of Lactobacillus casei confers protection to mice against the intracellular protozoan Babesia microti. Mice treated with L. casei orally or intraperitoneally were inoculated 7 days later with an infectious dose of B. microti. Mice treated with lactobacilli showed significant reduction in the percentage of parasitized erythrocytes (PPE) compared to untreated mice. When mice were inoculated intraperitoneally with L. casei 3 or 0 days before challenge with B. microti, the PPE was significantly lower compared to untreated mice and there were no differences between treated mice and mice immune to B. microti infection. When mice treated with live or dead L. casei were compared to mice inoculated with Freund Complete Adjuvant before a B. microti infection, a significant reduction of PPE was observed. These results show the protective effect of L. casei administered to mice against a B. microti infection and suggest that it might act by stimulating the innate immune system.
Lavin, S; Lastras, M E; Marco, I; Cabañes, F X
A case of fibrinopurulent bronchopneumonia associated with Moraxella bovis infection in a chamois (Rupicapra pyrenaica) is described. The animal, a 4-month-old female, was referred by the staff warden of the National Game Reserve of Freser-Setcases (Catalonia, north-eastern Spain). The animal was in good general condition and was found 4 h before death. On necropsy the lungs were congested and oedematous, with haemorrhagic areas in the cranial and middle lobes. The microscopic lesions were those of a fibrinopurulent bronchopneumonia. Microbiological study of the samples obtained showed numerous small beta-haemolytic colonies in pure culture, identified as Moraxella (Moraxella) bovis.
Hantsis-Zacharov, Elionora; Senderovich, Yigal; Halpern, Malka
Three Gram-negative, rod-shaped, oxidase-positive, aerobic, non-motile bacterial strains, designated H9T, H10 and H15, were isolated during a study on the diversity of culturable psychrotolerant bacteria in raw cow's milk. Comparisons of 16S rRNA gene sequences showed that the three strains were very closely related to each other (sequence similarities of 99.6-99.8 %). A polyphasic taxonomic study of the isolates resulted in their identification as members of the genus Chryseobacterium (family Flavobacteriaceae, phylum Bacteroidetes). The three strains showed < or =96.9 % sequence similarity with respect to the type strains of described Chryseobacterium species, indicating that H9T, H10 and H15 represent a novel species of the genus Chryseobacterium. The three strains grew at 7-37 degrees C (strain H10 grew at up to 41 degrees C), with 0-2.5 % NaCl and at pH 5.0-9.8. The dominant cellular fatty acids of strain H9T were 15 : 0 iso (38.9 %), 15 : 0 anteiso (15.6 %) and 17 : 0 iso 3-OH (12.7 %). Strain H10 also possessed 17 : 1 iso omega 9c (14.8 %) as a major fatty acid. On the basis of phenotypic properties and phylogenetic distinctiveness, the three milk isolates represent a novel species in the genus Chryseobacterium, for which the name Chryseobacterium bovis sp. nov. is proposed. The type strain is H9T (=LMG 24227T =DSM 19482T).
Hirata, Haruyuki; Kawai, Satoru; Maeda, Mari; Jinnai, Michio; Fujisawa, Kohei; Katakai, Yuko; Hikosaka, Kenji; Tanabe, Kazuyuki; Yasutomi, Yasuhiro; Ishihara, Chiaki
We demonstrate here the identification and phylogenetic characterization of Babesia microti (B. microti)-like parasite detected from a splenectomized Japanese macaque (Macaca fuscata fuscata) at a facility for laboratory animal science. On Day 133 after splenectomy, intra-erythrocytic parasites were found on light microscopic examination, and the level of parasitemia reached 0.3% on blood smear. Molecular characterization of the parasite using nested-polymerization chain reactions targeting the 18S rRNA, β-tubulin, and subunit 7 (eta) of the chaperonin-containing t-complex polypeptide 1 (CCT7) genes were identified as a B. microti-like parasite, designated the Japanese Macaque Babesia-1 (JM-1). PMID:21976563
Castro, Lyda R; Gabrielli, Simona; Iori, Albertina; Cancrini, Gabriella
The aim of the present study was to provide insight into the diversity of tick-borne pathogens circulating in Italy, carried/transmitted by Ixodes ricinus, one of the most abundant tick species in the country. A total of 447 specimens sampled in five areas of northeastern, central and insular Italy were analysed by polymerase chain reaction and sequencing for the presence of rickettsiae, borreliae and babesiae. Several rickettsial species of the spotted fever group of zoonotic concern and other zoonotic pathogens were found, such as Borrelia burgdorferi s.s., Borrelia afzelii, Borrelia garinii, and Babesia venatorum. These findings confirm a wide distribution of tick-borne bacterial and protozoan species in Italy, and highlight the sanitary importance of I. ricinus, often recorded as feeding on humans.
Yeruham, I; Hadani, A; Galker, F
Splenectomized gerbils (Acomys cahirinus) have been found to be susceptible to Babesia ovis applied via blood inoculation, whereas intact gerbils were not infected. Blood from splenectomized latently infected gerbils caused infection in a splenectomized lamb. Intact Acomys cahirinus gerbils did not become infected by B. ovis, and all captured rodents in an enzootic area, intact and splenectomized, were found to be negative for B. ovis. It can thus be concluded that Acomys cahirinus does not serve as a natural reservoir for B. ovis.
Anderson, J F; Johnson, R C; Magnarelli, L A; Hyde, F W; Myers, J E
Borrelia burgdorferi and Babesia microti were isolated from 35 of 51 white-footed mice (Peromyscus leucopus) and meadow voles (Microtus pennsylvanicus) captured on two Narragansett Bay, R.I., islands inhabited by deer, the principal host for the adult stages of the vector tick, Ixodes dammini. Immature ticks parasitized mice from both islands. From 105 mice captured on four other islands not inhabited by deer neither pathogen was isolated, nor were I. dammini found. PMID:3555339
Najm, Nour-Addeen; Meyer-Kayser, Elisabeth; Hoffmann, Lothar; Herb, Ingrid; Fensterer, Veronika; Pfister, Kurt; Silaghi, Cornelia
Wild canines which are closely related to dogs constitute a potential reservoir for haemoparasites by both hosting tick species that infest dogs and harbouring tick-transmitted canine haemoparasites. In this study, the prevalence of Babesia spp. and Theileria spp. was investigated in German red foxes (Vulpes vulpes) and their ticks. DNA extracts of 261 spleen samples and 1953 ticks included 4 tick species: Ixodes ricinus (n=870), I. canisuga (n=585), I. hexagonus (n=485), and Dermacentor reticulatus (n=13) were examined for the presence of Babesia/Theileria spp. by a conventional PCR targeting the 18S rRNA gene. One hundred twenty-one out of 261 foxes (46.4%) were PCR-positive. Out of them, 44 samples were sequenced, and all sequences had 100% similarity to Theileria annae. Similarly, sequencing was carried out for 65 out of 118 PCR-positive ticks. Theileria annae DNA was detected in 61.5% of the sequenced samples, Babesia microti DNA was found in 9.2%, and Babesia venatorum in 7.6% of the sequenced samples. The foxes were most positive in June and October, whereas the peak of tick positivity was in October. Furthermore, the positivity of the ticks was higher for I. canisuga in comparison to the other tick species and for nymphs in comparison to adults. The high prevalence of T. annae DNA in red foxes in this study suggests a reservoir function of those animals for T. annae. To our knowledge, this is the first report of T. annae in foxes from Germany as well as the first detection of T. annae and B. microti in the fox tick I. canisuga. Detection of DNA of T. annae and B. microti in three tick species collected from foxes adds new potential vectors for these two pathogens and suggests a potential role of the red fox in their natural endemic cycles.
Activity of rifapentine and its metabolite 25-O-desacetylrifapentine compared with rifampicin and rifabutin against Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis and M. bovis BCG.
Rastogi, N; Goh, K S; Berchel, M; Bryskier, A
The in vitro activity of rifapentine and its metabolite, 25-O:-desacetylrifapentine, as compared with that of rifampicin and rifabutin, was determined against Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis and M. bovis BCG. MICs were determined radiometrically and by the 1% proportional method using Middlebrook 7H11 agar. The bactericidal effect of the drugs was determined in parallel at selected concentrations. For drugsusceptible isolates of M. tuberculosis, the Bactec MICs of rifapentine and 25-O:-desacetylrifapentine were 0.03-0.06 mg/L and 0. 125-0.25 mg/L, respectively. Similar MICs were obtained for M. africanum (0.03-0.125 and 0.125-0.50 mg/L, respectively), and M. bovis (0.063-0.25 and 0.125-1.0 mg/L, respectively), but MICs were considerably lower for M. bovis BCG (0.008-0.063 mg/L for rifapentine and 0.016-0.125 mg/L for its metabolite). In general, MICs determined using 7H11 agar medium were usually one or two dilutions higher than those obtained using Bactec broth. When compared with rifampicin and rifabutin, the inhibitory activity of rifapentine for drug-susceptible isolates was roughly equal to that of rifabutin, and the inhibitory activity of 25-O:-desacetylrifapentine was comparable to that of rifampicin; however, rifapentine was somewhat more bactericidal than rifabutin at equal concentrations. Clinical isolates of M. tuberculosis with a high degree of resistance to rifampicin (MIC >/= 32 mg/L) were also highly resistant to rifabutin, rifapentine and 25-O:-desacetylrifapentine, although the MICs of rifabutin in this case were somewhat lower than the MICs of rifapentine.
Register, Karen B; Sacco, Randy E; Olsen, Steven C
Mycoplasma bovis has recently emerged as a significant and costly infectious disease problem in bison. A method for the detection of M. bovis-specific serum antibodies is needed in order to establish prevalence and transmission patterns. Enzyme-linked immunosorbent assays (ELISAs) validated for the detection of M. bovis-specific serum IgG in cattle are commercially available, but their suitability for bison sera has not been determined. A collection of bison sera, most from animals with a known history of infection or vaccination with M. bovis, was tested for M. bovis-specific IgG using commercially available kits as well as an in-house ELISA in which either cattle or bison M. bovis isolates were used as a source of antigen. Comparison of the results demonstrates that ELISAs optimized for cattle sera may not be optimal for the identification of bison seropositive for M. bovis, particularly those with low to moderate antibody levels. The reagent used for the detection of bison IgG and the source of the antigen affect the sensitivity of the assay. Optimal performance was obtained when the capture antigen was derived from bison isolates rather than cattle isolates and when a protein G conjugate rather than an anti-bovine IgG conjugate was used for the detection of bison IgG.
Tomlinson, A J; Chambers, M A; Wilson, G J; McDonald, R A; Delahay, R J
Heterogeneity in the progression of disease amongst individual wild animals may impact on both pathogen and host dynamics at the population level, through differential effects on transmission, mortality and reproductive output. The role of the European badger (Meles meles) as a reservoir host for Mycobacterium bovis infection in the UK and Ireland has been the focus of intense research for many years. Here, we investigate life-history correlates of infection in a high-density undisturbed badger population naturally infected with M. bovis. We found no evidence of a significant impact of M. bovis infection on female reproductive activity or success, with evidence of reproduction continuing successfully for several years in the face of M. bovis excretion. We also found evidence to support the hypothesis that female badgers are more resilient to established M. bovis infection than male badgers, with longer survival times following the detection of bacterial excretion. We discuss the importance of infectious breeding females in the persistence of M. bovis in badger populations, and how our findings in male badgers are consistent with testosterone-induced immunosuppression. In addition, we found significant weight loss in badgers with evidence of disseminated infection, based on the culture of M. bovis from body systems other than the respiratory tract. For females, there was a gradual loss of weight as infection progressed, whereas males only experienced substantial weight loss when infection had progressed to the point of dissemination. We discuss how these differences may be explained in terms of resource allocation and physiological trade-offs.
Ferreira, Daniele J; Zanine, Anderson M; Lana, Rogério P; Ribeiro, Marinaldo D; Alves, Guilherme R; Mantovani, Hilário C
The objective of the present study was to assess the chemical and bromatological composition and in situ degradability of elephant grass silages inoculated with Streptococcus bovis isolated from cattle rumen. A complete randomized design was used with four treatments and six replications: elephant grass silage, elephant grass silage inoculated with 10(6) CFU/g Streptococcus bovis JB1 strains; elephant grass silage inoculated with 106 CFU/g Streptococcus bovis HC5 strains; elephant grass silage inoculated with 106 CFU/g Enterococcus faecium with six replications each. The pH and ammoniacal nitrogen values were lower (P<0.05) for the silages inoculated with Streptococcus bovis JB1 and HC5, respectively. The silage inoculated with Streptococcus bovis had a higher crude protein content (P<0.05) and there were no differences for the fiber contents in the silage. The (a)soluble fraction degradability, especially in the silages inoculated with Streptococcus bovis JB1 and HC5, had higher values, 30.77 and 29.97%, for dry matter and 31.01 and 36.66% for crude protein, respectively. Inoculation with Streptococcus bovis improved the fermentation profile, protein value and rumen degradability of the nutrients.
Boothby, J T; Jasper, D E; Thomas, C B
Four cows were vaccinated with Mycoplasma bovis five times at two week intervals: three times subcutaneously in Freund's complete adjuvant, and two times with M. bovis alone in two of four quarters by intramammary infusion. The effect of vaccination on the immune response was evaluated in the serum and whey of the four vaccinated and control (placebo) cows experimentally challenged in two of four quarters with live M. bovis. Vaccination resulted in markedly increased M. bovis-specific, serum IgM, IgG and IgG2, but not IgA, reactivity. Challenge exposure with live M. bovis by intramammary infusion resulted in high specific serum IgM, IgG1 and IgG2 reactivity and a noticeable IgA response in both vaccinated and control cows. Whey from quarters on vaccinated cows had elevated, specific IgG1 reactivity at the time of challenge but no other differences were observed. Challenge exposure with live M. Bovis resulted in high antibody levels of all isotypes in quarters which were challenged, but highly elevated reactivities in unchallenged quarters occurred only with IgG1 and IgG2. These results indicate that vaccination elevated M. bovis-specific IgG1 but not other immunoglobulin reactivity in quarters on vaccinated cows, and that live organisms are necessary to elicit a local, specific IgA response. PMID:3567746
Background In many regions of the world, wild mammals act as reservoir of Mycobacterium bovis, a situation that prevents the eradication of bovine tuberculosis. In order to observe whether a strain isolated from a wild boar, previously tested as highly virulent in a mice model, is also virulent in cattle, we performed cattle experimental inoculation with this strain Results Groups of Friesian calves were either infected with the wild boar strain M. bovis 04-303 or with the bovine strain NCTC10772 as a control. We found that antigen-specific IFN-γ release in whole blood samples occurred earlier in animals infected with M. bovis 04-303. Both M. bovis strains resulted in a positive skin test, with animals infected with the wild boar isolate showing a stronger response. These results and the presence of more severe organ lesions, with granuloma and pneumonic areas in cattle demonstrate that the wild boar isolate is more virulent than the NCTC10772 strain. Additionally, we tested the infectivity of the M. bovis strains in guinea pigs and found that M. bovis 04-303 had the highest pathogenicity. Conclusions M. bovis strains isolated from wild boars may be pathogenic for cattle, producing TB lesions. PMID:21745408
Tomassone, L; Pagani, P; De Meneghi, D
A reverse line blot hybridisation (RLB) assay was applied to screen Amblyomma variegatum adult ticks (n = 504) collected from N'Dama cattle in the Republic of Guinea. In a PCR, the V1 hypervariable region of the 16S ribosomal RNA (rRNA) gene was amplified with a set of primers unique for species of the genera Anaplasma and Ehrlichia, and the V4 hypervariable region of the 18S rRNA gene was amplified with primers specific for members of the genera Theileria and Babesia. Amplified PCR products from A. variegatum ticks were hybridised onto a membrane, to which oligonucleotide probes species-specific for Ehrlichia/Anaplasma and Theileria/Babesia parasites were covalently linked. No pathogens belonging to Ehrlichia/Anaplasma species were found, while 10 DNA samples resulted positive for Babesia caballi and 5 samples for Theileria velifera. This is the first report of B. caballi in A. variegatum ticks. One of the B. caballi positive samples was sequenced. This new strain (BcabGuinea) showed a 97% similarity to the Z15104 B. caballi GenBank sequence.
Munang'andu, Hetron Mweemba; Munyeme, Musso; Nambota, Andrew Mubila; Nalubamba, King Shimumbo; Siamudaala, Victor M
Babesia spp. were detected from 4 asymptomatic pukus captured on a game ranch in central Zambia in October 2008. Blood smears were examined in 4 species of aymptomatic free-ranging antelopes, namely the puku (Kobus vordanii), reedbuck (Redunca arundinum), bushbuck (Tragelaphus sylvaticus), and kudu (Tragelaphus strepsiceros), and showed the presence of Babesia parasites only in the puku. In the puku, the prevalence of babesiosis was estimated at 33.3% (n = 12), while the overall prevalence in all examined animals was 8.5% (n = 47). The parasites showed morphological characteristics of paired ring-like stages with the length varying between 1.61 µm and 3.02 µm (mean = 2.12 µm, n = 27; SD = 0.76 µm). Both the infected and non-infected pukus showed good body condition scores (BCS), while the dominant tick species detected from all animals were Rhipicephalus appendiculatus, Rhipicephalus spp., and Boophilus spp. To our knowledge this is the first report of Babesia spp. infection in pukus in Zambia. These findings suggest that wildlife could play an important role in the epidemiology of babesiosis in Zambia.
Rotondano, Tereza Emmanuelle de Farias; Almeida, Herta Karyanne Araújo; Krawczak, Felipe da Silva; Santana, Vanessa Lira; Vidal, Ivana Fernandes; Labruna, Marcelo Bahia; de Azevedo, Sérgio Santos; Ade lmeida, Alzira Maria Paiva; de Melo, Marcia Almeida
This study assessed the occurrence of Ehrlichia spp., Babesia spp. and Hepatozoon spp. infections in 100 tick-harboring dogs from a semiarid region of the State of Paraíba, Northeastern Brazil. Blood samples and ticks were collected from the animals, and a questionnaire was submitted to dog owners to obtain general data. Blood samples were used to perform hemogram, direct blood smear and immunological and molecular hemoparasite detection. The 1,151 ticks collected were identified as Rhipicephalus sanguineus; direct smears revealed E. canis-like morulae in the monocytes of 4% (4/100) of the non-vaccinated female dogs, and 34% and 25% of the dogs tested positive for Ehrlichia canis by indirect immunofluorescence assay (IFA) and polymerase chain reaction (PCR), respectively. Blood smear examination revealed Babesia-suggestive merozoites in the erythrocytes of 2% (2/100) of the animals. Babesia vogeli was detected by PCR in ten animals (10%) and was correlated with young age (p = 0.007) and thrombocytopenia (p = 0.01). None of the animals showed Hepatozoon spp. positivity. These results indicate that E. canis is the main tick-borne canine pathogen in the study area and provide the first report of B. vogeli infection in dogs from Paraiba State.
Corredoira, J C; Alonso, M P; García, J F; Casariego, E; Coira, A; Rodriguez, A; Pita, J; Louzao, C; Pombo, B; López, M J; Varela, J
The aim of this study was to determine the clinical significance of Streptococcus salivarius isolates recovered from blood cultures and compare them with isolates of Streptococcus bovis biotypes I and II. Seventeen of the 52 (32%) S. salivarius isolates recovered were considered clinically significant, compared with 62 of the 64 (97%) S. bovis isolates (p<0.0001). Bacteremia caused by S. salivarius occurred mostly in patients who showed relevant disruption of the mucous membranes and/or serious underlying diseases. Patients with S. salivarius bacteremia were younger than those with S. bovis bacteremia (57 vs. 67 years; p<0.01). Patients with S. salivarius bacteremia and patients with S. bovis II bacteremia had similar rates of endocarditis, colon tumors, and non-colon cancer. On the other hand, when compared with S. bovis I bacteremia, S. salivarius bacteremia was associated with lower rates of endocarditis (18% vs. 74%, respectively) (p<0.01) and colon tumors (0% vs. 57%, respectively) (p<0.005) and higher rates of non-colon cancer (53% vs. 9.5%, respectively) (p<0.01). Bacteremia caused by S. bovis II had a hepatobiliary origin in 50% of the patients, while, in contrast, that due to S. salivarius or S. bovis I was less frequently associated with a hepatobiliary origin (12% and 5%, respectively) (p<0.00001). The rate of penicillin resistance was 31% among S. salivarius isolates and 0% among S. bovis isolates (p<0.0001). In conclusion, the clinical characteristics of S. salivarius bacteremia and S. bovis II bacteremia are similar, and the isolation of S. salivarius in blood should not be systematically regarded as contamination.
The causative agent of tuberculosis (TB) in cattle is Mycobacterium bovis. The characteristic lesions of bovine TB are well-organized pulmonary granulomas. Gamma delta T cells are a unique subset of nonconventional T cells that play major roles in both the innate and adaptive arms of the immune sys...
We report the draft genome sequences of Streptococcus bovis type strain ATTC 33317 (CVM42251) isolated from cow dung and strain JB1 (CVM42252) isolated from a cow rumen in 1977. Strains were subjected to Next Generation sequencing and the genome sizes are approximately 2 MB and 2.2 MB, respectively....
The objective of this study was to identify microRNAs associated with a serum antibody response to Mycoplasma bovis in beef cattle. Serum from sixteen beef calves was collected at three points: in summer after calves were born, in fall at weaning, and in the following spring. All sera collected in t...
We recently reported that on 14 dairy farms from Vermont to Florida ~85% of pre-weaned dairy calves were infected with zoonotic Cryptosporidium parvum whereas only 1-2% of post-weaned calves and 1-2 year-old heifers were infected with this species. Cryptosporidium bovis and the deer-like genotype w...
Lesellier, Sandrine; Corner, Leigh; Costello, Eamon; Sleeman, Paddy; Lyashchenko, Konstantin P; Greenwald, Rena; Esfandiari, Javan; Glyn Hewinson, R; Chambers, Mark; Gormley, Eamonn
The Eurasian badger (Meles meles) is a wildlife reservoir for Mycobacterium bovis infection in Ireland and Great Britain and has been implicated in the transmission of tuberculosis to cattle. Vaccination of badgers is an option that could be used as part of a strategy to control the disease. In this study we used an endobronchial infection procedure to inoculate groups of badgers with three different doses (3x10(3), 2x10(2) and <10 Colony Forming Units (CFUs)) of M. bovis. After 17 weeks the disease status of each animal was determined by post-mortem pathology and culture for M. bovis. Each of the inoculum doses resulted in establishment of infection in the badgers. The cell-mediated immune (CMI) responses were measured by lymphocyte transformation assay (LTA) of peripheral blood mononuclear cells (PBMCs) cultured with bovine tuberculin (PPD-B). In each infected group the CMI responses increased with a kinetic profile corresponding to the delivered dose and the post-mortem pathology. The serological responses were measured by ELISA and a multi-antigen print immunoassay (MAPIA) in order to investigate any changes in the antigenic repertoire associated with different infective doses. In contrast to the CMI responses, the ELISA and MAPIA showed that the recognition of antigens by the badgers was intermittent and not strongly influenced by the dose of M. bovis.
Balseiro, Ana; González-Quirós, Pablo; Rodríguez, Óscar; Francisca Copano, M; Merediz, Isabel; de Juan, Lucía; Chambers, Mark A; Delahay, Richard J; Marreros, Nelson; Royo, Luis J; Bezos, Javier; Prieto, José M; Gortázar, Christian
Recent studies suggest that badgers may be a potential reservoir of Mycobacterium bovis infection for cattle in Northern Spain. The objective of this study was to investigate potential epidemiological links between cattle and badgers. Culture and molecular typing data were available for cattle culled during the national tuberculosis (TB) eradication campaigns between 2008 and 2012, as well as from 171 necropsied badgers and 60 live animals trapped and examined over the same time period. Mycobacterium tuberculosis complex strains were isolated from pooled tissues of 14 (8.2%) necropsied badgers, of which 11 were identified as M. bovis: six different spoligotypes of M. bovis were subsequently identified. In two geographical locations where these isolates were shared between cattle and badgers, infected cattle herds and badgers lived in close contact. Although it remains unclear if badgers are a maintenance or spill-over host of M. bovis in this setting, it would appear prudent to have precautionary measures in place to reduce contact between cattle and badgers.
Byrne, W; Markey, B; McCormack, R; Egan, J; Ball, H; Sachse, K
To study the course of clinical mycoplasma mastitis and investigate its potential for persistence, 10(8) colony-forming units (cfu) of an Irish isolate of Mycoplasma bovis was inoculated aseptically into the right fore teat canal of three lactating cows. M bovis rapidly colonised the infected quarters and grew exponentially to more than 10(10) cfu/ml within the first three days, and spread to other quarters of each of the three cows within five to 10 days. After periods of between 24 and 72 hours the infected quarters became distended and sensitive to touch, and their secretions changed from containing visible particles, to a seropurulent exudate, to an aqueous suspension of fine particles which formed a sediment after a sample was collected. M bovis-specific antibody levels increased to varying degrees in all three cows. Subsequently, the concentrations of mycoplasma decreased to less than 10(7) cfu/ml in two of the cows, but remained at more than 10(8) cfu/ml to the end of the lactation of the other cow. Apparently normal milk was secreted by one of the cows within a month of the challenge, and by the other two cows at the start of their next lactation. However, in two of the cows subclinical M bovis infection persisted through the dry periods and into their next lactations.
In 1994, white-tailed deer in northeast Michigan were found to be harboring Mycobacterium bovis, the causative agent of tuberculosis in most animals including humans. Although deer likely contracted tuberculosis from cattle in the early 20th century, when the disease was present in Michigan cattle, ...
The presence of tuberculosis due to Mycobacterium bovis in captive and free-ranging wildlife remains one of the greatest challenges to eradication of tuberculosis in the United States. A possible addition to current control measures could be vaccination of deer to prevent infection, disease, or tran...
King, Hayley C; Murphy, Andrew; James, Phillip; Travis, Emma; Porter, David; Sawyer, Jason; Cork, Jennifer; Delahay, Richard J; Gaze, William; Courtenay, Orin; Wellington, Elizabeth M
The incidence of Mycobacterium bovis, the causative agent of bovine tuberculosis, in cattle herds in the United Kingdom is increasing, resulting in substantial economic losses. The European badger (Meles meles) is implicated as a wildlife reservoir and is the subject of control measures aimed at reducing the incidence of infection in cattle populations. Understanding the epidemiology of M. bovis in badger populations is essential for directing control interventions and understanding disease spread; however, accurate diagnosis in live animals is challenging and currently uses invasive methods. Here we present a noninvasive diagnostic procedure and sampling regimen using field sampling of latrines and detection of M. bovis with quantitative PCR tests, the results of which strongly correlate with the results of immunoassays in the field at the social group level. This method allows M. bovis infections in badger populations to be monitored without trapping and provides additional information on the quantities of bacterial DNA shed. Therefore, our approach may provide valuable insights into the epidemiology of bovine tuberculosis in badger populations and inform disease control interventions.
Fitzgerald, Scott D; Zwick, Laura S; Diegel, Kelly L; Berry, Dale E; Church, Steven V; Sikarskie, James G; Kaneene, John B; Reed, Willie M
The goal of this study was to evaluate the susceptibility of North American opossums (Didelphis virginiana) to aerosol inoculation of Mycobacterium bovis at two dose levels in order to gain information on disease pathogenesis, fecal shedding of the organism, and the potential role that opossums play in the spread of this disease in nature. Six opossums received high dose (1 x 10(7) colony forming units (cfu) by aerosol inoculation, six opossums received low dose (1 x 10(3) cfu inoculation, and six opossums were sham-inoculated with sterile water and served as controls. Lungs were the most frequently infected tissues, with nine of 12 inoculated opossums positive for M. bovis on culture. Gross lesions consisted of multifocal pneumonia and enlarged lymph nodes. Microscopically, granulomatous pneumonia and granulomatous lymphadenitis associated with acid-fast bacilli were present in eight of 12 inoculated opossums. Fecal shedding of M. bovis was uncommon at both inoculation doses. While opossums were highly susceptible to aerosol inoculation of M. bovis, they did not become emaciated or develop widely disseminated lesions. From this study, opossums may transmit tuberculosis by aerosol infection to other opossums in close contact and serve as a source of infection to carnivores that feed upon them, however, transmission of the disease to large herbivores by fecal shedding or direct contact may be less likely.
Complex and dynamic interactions involving domestic animals, wildlife and humans create environments favorable to the emergence of new diseases, or re-emergence of diseases in new host species. Today, reservoirs of Mycobacterium bovis, the causative agent of tuberculosis in animals and a serious zoo...
Background Over the past two decades, highly virulent strains of Mycobacterium tuberculosis have emerged and spread rapidly in humans, suggesting a selective advantage based upon virulence. A similar scenario has not been described for Mycobacterium bovis infection in cattle (i.e., Bovine Tuberculos...
Buss, Peter E.; van Helden, Paul D.; Parsons, Sven D.C.
In 2016, an emaciated black rhinoceros (Diceros bicornis) was found in Kruger National Park, South Africa. An interferon-γ response was detected against mycobacterial antigens, and lung tissue was positive for Mycobacterium bovis. This case highlights the risk that tuberculosis presents to rhinoceros in M. bovis–endemic areas. PMID:28221132
Fructans from pasture can be fermented by Gram-positive bacteria (e.g., Streptococcus bovis) in the equine hindgut, increasing production of lactic acid and decreasing pH. The degree of polymerization (DP) of fructans has been suggested to influence fermentation rates. The objective of the current ...
Background. Mycoplasma bovis is a cause of respiratory disease in cattle and the bacterium most frequently isolated from bovine respiratory disease complex. It has recently emerged as a major health problem in bison, causing pharyngitis, pneumonia, arthritis, dystocia and abortion. In cattle, M. b...
Giovannini, S; Zanoni, M G; Salogni, C; Cinotti, S; Alborali, G L
Mycoplasma bovis is an important cause of bovine respiratory disease, especially in young calves where it can also cause arthritis, tenosynovitis and otitis. During 2009 and 2010 a survey was carried out on carcasses of calves less than one month old sent to the Diagnostic Laboratory of the Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna in Brescia, regardless of the presence of lung lesions, to detect this pathogen. PCR tests for Mycoplasma spp. and M. bovis were applied. 83 out of 224 (37%) lung tissue samples examined were positive at PCR test for Mycoplasma spp.; in 64 cases of these we observed typical respiratory lesions (P<0.001). M. bovis was identified in 26 out of 83 (31%) lung tissue samples positive at PCR test for Mycoplasma spp.; in 24 cases of these we observed typical respiratory lesions (P=0.039). Our data demonstrate that presence of Mycoplasma spp. and M. bovis positively correlates with pneumonic lung lesions in young dairy calves.
Previous work in small animal laboratory models of tuberculosis have shown that vaccination strategies based on heterologous prime-boost protocols using Mycobacterium bovis bacille Calmette-Guerin (BCG) to prime and Modified Vaccinia Ankara strain (MVA85A) or recombinant attenuated adenoviruses (Ad8...
Young, Sarah L; Slobbe, Lynn J; Peacey, Matthew; Gilbert, Sarah C; Buddle, Bryce M; de Lisle, Geoffrey W; Buchan, Glenn S
DNA-based vaccines, alone or in combination with other sub-unit vaccination regimes, represent an alternative to live mycobacterial vaccines for protective immunization against tuberculosis. Here, we have used a murine immunization or Mycobacterium bovis aerosol challenge model to assess the immunogenicity and protective efficacy of mycobacterial DNA vaccines. Mice that received immunization with DNA constructs encoding M. bovis antigen 85A (Ag85-A) and arget(ESAT-6) produced measurable interferon-gamma (IFN-gamma) responses to CD4(+) T-cell epitope-peptide recall antigens in vitro. The magnitude of these responses was enhanced by co-delivery of a construct encoding murine cytokines (macrophage inhibitory protein (MIP)-1 alpha or interleukin(IL)-7), although they did not the match responses observed in mice that received Bacille Calmette-Guerin(BCG) immunisation. In contrast, DNA priming followed by boosting with modified vaccinia Ankara (MVA) vaccine (expressing M. tuberculosis Ag85-A) invoked higher IFN-gamma levels, with the most immunogenic regime of Ag85 or ESAT or IL-7 prime followed by MVA boost being of commensurate immunogenicity to BCG. Despite this, neither DNA alone nor DNA-prime or MVA boost regimes conferred measurable protection against aerosol challenge with virulent M. bovis. These data highlight both the promise and the shortcomings of new generation subunit tuberculosis vaccines, with particular emphasis on their potential as vaccines against M. bovis.
Cardoso, M A; Cardoso, R F; Hirata, R D C; Hirata, M H; Leite, C Q F; Santos, A C B; Siqueira, V L D; Okano, W; Rocha, N S; Lonardoni, M V C
Thirty-five lymph node samples were taken from animals with macroscopic lesions consistent with Mycobacterium bovis infection. The animals were identified by postmortem examination in an abattoir in the northwestern region of state of Paraná, Brazil. Twenty-two of the animals had previously been found to be tuberculin skin test positive. Tissue samples were decontaminated by Petroff's method and processed for acid-fast bacilli staining, culture in Stonebrink and Lowenstein-Jensen media and DNA extraction. Lymph node DNA samples were amplified by PCR in the absence and presence (inhibitor controls) of DNA extracted from M. bovis culture. Mycobacterium bovis was identified in 14 (42.4%) lymph node samples by both PCR and by culture. The frequency of PCR-positive results (54.5%) was similar to that of culture-positive results (51.5%, P > 0.05). The percentage of PCR-positive lymph nodes increased from 39.4% (13/33) to 54.5% (18/33) when samples that were initially PCR-negative were reanalysed using 2.5 microl DNA (two samples) and 1 : 2 diluted DNA (three samples). PCR sensitivity was affected by inhibitors and by the amount of DNA in the clinical samples. Our results indicate that direct detection of M. bovis in lymph nodes by PCR may be a fast and useful tool for bovine tuberculosis epidemic management in the region.
Romero, R E; Garzón, D L; Mejía, G A; Monroy, W; Patarroyo, M E; Murillo, L A
Tuberculosis, caused by Mycobacterium bovis is emerging as the most important disease affecting cattle. Furthermore, it results in a major public health problem when transmitted to humans. Due to its difficult and non-specific diagnosis, M. bovis has been declared to be one of the etiologic agents causing significant economic loss in the cattle industry. Our group evaluated a more rapid and specific method, based on a new polymerase chain reaction species-specific primers, which amplifies a 470-base pair fragment of the M. bovis genome. A total of 275 milk-producing cows were studied by intradermal tuberculin test (ITT) which gave 184 positive and 91 negative cases. From them, 50 animals were taken from a cattle ranch free of tuberculosis. Three different samples were collected from each animal (blood, nasal mucus, and milk). Positive results were obtained from 26 animals by PCR (11.4%), 1 by bacteriological culturing (0.4%) and 1 by bacilloscopy (0.4%). This finding suggests, as in previous reports, that ITT, normally used for bovine tuberculosis detection, has the inconvenience of having a broad range of specificity and sensitivity, and the PCR technique is a more specific and sensitive test to detect infection associated with M. bovis. Therefore, we propose this PCR assay as a useful tool in the epidemiological characterization of infected animals in areas considered to be at high risk of transmission. Images Figure 1. PMID:10369566
Milián-Suazo, Feliciano; Pérez-Guerrero, Laura; Arriaga-Díaz, Camila; Escartín-Chávez, Minerva
The purpose of this study was to evaluate the role of Mycobacterium bovis in human cases of tuberculosis (TB) in an endemic area of the disease in cattle. Sputum, urine and other tissue samples were obtained from: (1) TB-symptomatic patients, (2) dairy farm workers and (3) abattoir workers. Samples of macroscopic lesions suspicious of TB were also obtained from cattle at slaughter in the same geographic area. A total of 562 human samples were collected: 255 from symptomatic patients, 218 from farm workers and 93 from abattoir workers. Samples were analysed by the bacillus acido-alcohol resitant (BAAR) and polymerase chain reaction (PCR) tests and cultured in Stonebrink and Löwenstein-Jensen. Spoligotyping was performed in all isolates obtained by culture and the DNA obtained by PCR. From the total number of human cases, 34 (6%) showed M. bovis spoligotype; eight spoligotypes from cattle showed an identical pattern to three spoligotypes from humans; a different set of spoligotypes from cattle (n = 8) had only one spacer difference to a set of spoligotypes from humans (n = 2). These results provide further evidence that infected cattle represent a risk to public health and support previous reports about the role of M. bovis in Mexican patients. There is no doubt that genotyping M. bovis isolates collected from cattle may have a substantial impact on our understanding of the epidemiology of TB.
Amram, E.; Mikula, I.; Schnee, C.; Ayling, R. D.; Nicholas, R. A. J.; Rosales, R. S.; Harrus, S.
Mycoplasma bovis isolates with decreased susceptibilities to tetracyclines are increasingly reported worldwide. The acquired molecular mechanisms associated with this phenomenon were investigated in 70 clinical isolates of M. bovis. Sequence analysis of the two 16S rRNA-encoding genes (rrs3 and rrs4 alleles) containing the primary binding pocket for tetracycline (Tet-1 site) was performed on isolates with tetracycline hydrochloride MICs of 0.125 to 16 μg/ml. Mutations at positions A965T, A967T/C (Escherichia coli numbering) of helix 31, U1199C of helix 34, and G1058A/C were identified. Decreased susceptibilities to tetracycline (MICs, ≥2 μg/ml) were associated with mutations present at two (A965 and A967) or three positions (A965, A967, and G1058) of the two rrs alleles. No tet(M), tet(O), or tet(L) determinants were found in the genome of any of the 70 M. bovis isolates. The data presented correlate (P < 0.0001) the mutations identified in the Tet-1 site of clinical isolates of M. bovis with decreased susceptibility to tetracycline. PMID:25403668
Wibberg, Daniel; Winkler, Anika; Straube, Eberhard; Karrasch, Matthias; Keller, Peter M.
Here, we present the draft genome sequence of Mycobacterium bovis BCG S4-Jena, a tuberculosis vaccine strain. The genome of S4-Jena is represented by 48 scaffolds, consisting of 132 scaffolded contigs and amounting to a size of about 4.2 Mb. New genes potentially encoding a phage fragment were identified in the genome. PMID:27103721
Viljoen, Ignatius M; van Helden, Paul D; Millar, Robert P
Mycobacterium bovis has global public-health and socio-economic significance and can infect a wide range of species including the lion (Panthera leo) resulting in tuberculosis. Lions are classified as vulnerable under the IUCN Red List of Threatened Species and have experienced a 30% population decline in the past two decades. However, no attempt has been made to collate and critically evaluate the available knowledge of M. bovis infections in lions and potential effects on population. In this review we set out to redress this. Arguments suggesting that ingestion of infected prey animals are the main route of infection for lions have not been scientifically proven and research is needed into other possible sources and routes of infection. The paucity of knowledge on host susceptibility, transmission directions and therefore host status, manifestation of pathology, and epidemiology of the disease in lions also needs to be addressed. Advances have been made in diagnosing the presence of M. bovis in lions. However, these diagnostic tests are unable to differentiate between exposure, presence of infection, or stage of disease. Furthermore, there are contradictory reports on the effects of M. bovis on lion populations with more data needed on disease dynamics versus the lion population's reproductive dynamics. Knowledge on disease effects on the lion reproduction and how additional stressors such as drought or co-morbidities may interact with tuberculosis is also lacking. Filling these knowledge gaps will contribute to the understanding of mycobacterial infections and disease in captive and wild lions and assist in lion conservation endeavours.
Hermosilla, Carlos; Zahner, Horst; Taubert, Anja
Eimeria bovis is an important coccidian parasite of cattle causing severe diarrhea in young animals. Its first schizogony takes place in endothelial cells of the ileum resulting in the formation of macroschizonts 14-18 days p.i. This longlasting development suggests a particular immune evasion strategy of the parasite. Here, we analyse early innate immune reactions to E. bovis by determining the adhesion of polymorphonuclear neutrophils (PMN) to infected endothelial cell layers under flow conditions and the transcription of adhesion molecule genes in infected host cells. Bovine umbilical vein endothelial cells (BUVEC) were infected with E. bovis sporozoites. Sporozoites invaded BUVEC within 1h and the first mature macroschizonts occurred 14 days p.i. PMN adhesion was enhanced in E. bovis-infected BUVEC layers as early as 8h p.i.; maximum adhesion occurred 48 h p.i. Increased adhesion rates persisted until the end of the observation period at 14 days p.i. PMN adhered to both infected and uninfected cells within monolayers, suggesting paracrine cell activation. E. bovis infection upregulated the transcription of genes encoding for P-selectin, E-selectin, vascular cellular adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1). Most marked effects concerned E-selectin followed by P-selectin, VCAM-1 and ICAM-1. Increased transcript levels were found beginning 30 min p.i. and maximum values occurred 1-2h p.i. (P-selectin) and 2-4h p.i. (E-selectin, VCAM-1, ICAM-1). By 12-24h p.i. levels had decreased to those of uninfected controls. Tumor necrosis factor alpha (TNFalpha)-induced PMN adhesion was significantly reduced in infected vs. uninfected BUVEC. Eimeria bovis also had suppressive effects on TNFalpha-mediated upregulation of adhesion molecule gene transcription. The data presented here suggest that infection of BUVEC with E. bovis on one hand induces proinflammatory reactions resulting in enhanced PMN adhesion mediated by upregulated adhesion
Background Bovine tuberculosis (TB) caused by Mycobacterium bovis is primarily a disease of ruminants, particularly cattle (Bos primigenius) and buffalo (Syncerus caffer), and is endemic in most developing countries. To date, studies done in Uganda have documented the prevalence of M. bovis in cattle, humans and wild life, in addition to non-tuberculous mycobacteria in pigs. Pigs are increasingly becoming an important component of the livestock sector and share the human ecosystem in rural Uganda. It is therefore of public health interest that they are not a source of human infections. As a follow up to previously published findings on mycobacteria in pigs, this study was aimed at investigating the occurrence and molecular characteristics of M. bovis detected in slaughter pigs in Mubende district, Uganda. One hundred fifty mesenteric lymph nodes with lesions suggestive of mycobacterial infections were collected from approximately one thousand slaughtered pigs in Mubende district over a period of five months. The isolation and identification of M. bovis was done using conventional mycobacteriological methods. Mycobacteria belonging to the Mycobacterium tuberculosis complex (MTC) were identified to species level using deletion analysis. Molecular typing was done using Spoligotyping and MIRU-VNTR analysis. Molecular data were analysed and interpreted using MIRU-VNTR plus, SpolDB4.0 and the Mycobacterium bovis spoligo database. Results Of the examined animals, one boar and two sows from Madudu Sub County were infected with M. bovis which presented as lesions of a deep yellow colour and a grit-like texture in the mesenteric lymph nodes. This represents 2% (3/150) of the lymph nodes where lesions suggestive of mycobacterial infections were detected. Molecular analysis revealed that the isolates from the infected pigs showed identical MIRU-VNTR profile and spoligotype (SB1469). Conclusions This is the first study documenting the occurrence of M. bovis in slaughter pigs in
A worldwide standard on food irradiation was adopted in 1983 by Codex Alimentarius Commission of the Joint Food Standard Programme of the Food and Agriculture Organization (FAO) of the United Nations and the World Health Organization (WHO). As a result, 41 countries have approved the use of irradiation for treating one or more food items and the number is increasing. Generally, irradiation is used to: food loses, food spoilage, disinfestation, safety and hygiene. The number of countries which use irradiation for processing food for commercial purposes has been increasing steadily from 19 in 1987 to 33 today. In the frames of the national programme on the application of irradiation for food preservation and hygienization an experimental plant for electron beam processing has been established in Institute of Nuclear Chemistry and Technology. The plant is equipped with a small research accelerator Pilot (19MeV, 1 kW) and an industrial unit Elektronika (10MeV, 10 kW). On the basis of the research there were performed at different scientific institutions in Poland, health authorities have issued permission for irradiation for: spices, garlic, onions, mushrooms, potatoes, dry mushrooms and vegetables.
Hungate, F.P.; Riemath, W.F.; Bunnell, L.R.
A tissue irradiator is provided for the in-vivo irradiation of body tissue. The irradiator comprises a radiation source material contained and completely encapsulated within vitreous carbon. An embodiment for use as an in- vivo blood irradiator comprises a cylindrical body having an axial bore therethrough. A radioisotope is contained within a first portion of vitreous carbon cylindrically surrounding the axial bore, and a containment portion of vitreous carbon surrounds the radioisotope containing portion, the two portions of vitreous carbon being integrally formed as a single unit. Connecting means are provided at each end of the cylindrical body to permit connections to blood- carrying vessels and to provide for passage of blood through the bore. In a preferred embodiment, the radioisotope is thulium-170 which is present in the irradiator in the form of thulium oxide. A method of producing the preferred blood irradiator is also provided, whereby nonradioactive thulium-169 is dispersed within a polyfurfuryl alcohol resin which is carbonized and fired to form the integral vitreous carbon body and the device is activated by neutron bombardment of the thulium-169 to produce the beta-emitting thulium-170.
Müller, Borna; Steiner, Benjamin; Bonfoh, Bassirou; Fané, Adama; Smith, Noel H; Zinsstag, Jakob
Background Mali is one of the most important livestock producers of the Sahel region of Africa. A high frequency of bovine tuberculosis (BTB) has been reported but surveillance and control schemes are restricted to abattoir inspections only. The objective of this study was to conduct, for the first time, molecular characterisation of Mycobacterium bovis strains isolated from cattle slaughtered at the Bamako abattoir. Of 3330 animals screened only 60 exhibited gross visible lesions. From these animals, twenty strains of M. bovis were isolated and characterised by spoligotyping. Results Organ lesions typical of BTB were most often detected in the liver, followed by the lung and the peritoneum. M. bovis was isolated from 20 animals and 7 different spoligotypes were observed among these 20 strains; three of the patterns had not been previously reported. Spoligotype patterns from thirteen of the strains lacked spacer 30, a characteristic common in strains of M. bovis found in Chad, Cameroon and Nigeria. However, unlike the other three Central African countries, the majority of spoligotype patterns observed in Mali also lacked spacer 6. Of the remaining seven strains, six had spoligotype patterns identical to strains commonly isolated in France and Spain. Conclusion Two groups of M. bovis were detected in cattle slaughtered at the Bamako abattoir. The spoligotype pattern of the first group has similarities to strains previously observed in Chad, Cameroon and Nigeria. The additional absence of spacer 6 in the majority of these strains suggests a Mali specific clone. The spoligotype patterns of the remaining strains suggest that they may have been of European origin. PMID:18637160
Fend, R; Geddes, R; Lesellier, S; Vordermeier, H-M; Corner, L A L; Gormley, E; Costello, E; Hewinson, R G; Marlin, D J; Woodman, A C; Chambers, M A
It is estimated that more than 50 million cattle are infected with Mycobacterium bovis worldwide, resulting in severe economic losses. Current diagnosis of tuberculosis (TB) in cattle relies on tuberculin skin testing, and when combined with the slaughter of test-positive animals, it has significantly reduced the incidence of bovine TB. The failure to eradicate bovine TB in Great Britain has been attributed in part to a reservoir of the infection in badgers (Meles meles). Accurate and reliable diagnosis of infection is the cornerstone of TB control. Bacteriological diagnosis has these characteristics, but only with samples collected postmortem. Unlike significant wild animal reservoirs of M. bovis that are considered pests in other countries, such as the brushtail possum (Trichosurus vulpecula) in New Zealand, the badger and its sett are protected under United Kingdom legislation (The Protection of Badgers Act 1992). Therefore, an accurate in vitro test for badgers is needed urgently to determine the extent of the reservoir of infection cheaply and without destroying badgers. For cattle, a rapid on-farm test to complement the existing tests (the skin test and gamma interferon assay) would be highly desirable. To this end, we have investigated the potential of an electronic nose (EN) to diagnose infection of cattle or badgers with M. bovis, using a serum sample. Samples were obtained from both experimentally infected badgers and cattle, as well as naturally infected badgers. Without exception, the EN was able to discriminate infected animals from controls as early as 3 weeks after infection with M. bovis, the earliest time point examined postchallenge. The EN approach described here is a straightforward alternative to conventional methods of TB diagnosis, and it offers considerable potential as a sensitive, rapid, and cost-effective means of diagnosing M. bovis infection in cattle and badgers.
McCluskey, B; Lombard, J; Strunk, S; Nelson, D; Robbe-Austerman, S; Naugle, A; Edmondson, A
From 2002 to 2013, bovine tuberculosis (bTB) caused by Mycobacterium bovis (M. bovis) has been detected on numerous dairies in California. In total, twelve herds had bTB detected and are included in the case series which describes these recent outbreaks and discusses potential pathways of introduction. Epidemiological investigations to determine the initial source of bTB in each herd included obtaining data on likely pathways of pathogen introduction. Pathways included purchasing cattle, use of heifer-raising operations, commingling of cattle at greater risk of exposure to infected cattle with cattle destined for California dairies, contact with infected wildlife, exposure to humans with bTB infections, community and neighboring herds and others. Epidemiologic and molecular typing data confirmed the source of infection in 3 herds and probable sources of infection in 2 herds. In the 7 remaining herds described in this case series an epidemiologic link to a source could not be determined and molecular typing results did not associate M. bovis isolates acquired from these herds with another specific U.S. herd or U.S.-born animal. Preventing new introductions of M. bovis onto California dairies will require rigorous epidemiologic investigation of all the potential pathways of introduction discussed here. The root cause(s) of bTB on California dairies is certainly multifactorial with complex interactions of herd management practices, importation of cattle at greater risk of exposure to infected cattle, and the potential of human M. bovis exposure. The extensive use of molecular typing has improved epidemiologists' ability to narrow the scope of potential sources.
Lutz, Kathleen; Schmitt, Sigrid; Linder, Monica; Hermosilla, Carlos; Zahner, Horst; Taubert, Anja
The proteome of Eimeria bovis meront I-carrying host cells was analyzed by two-dimensional gel electrophoresis (2DE) at 14 days p.i. and compared to non-infected control cells. A total of 221 protein spots were modulated in their abundance in E. bovis-infected host cells and were subsequently analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectometry (MALDI-TOF-MS). These analyses identified 104 proteins in total with 25 host cell proteins being up-regulated and 79 proteins being down-regulated in E. bovis-infected host cells. Moreover, 20 newly expressed proteins were identified exclusively in E. bovis-infected host cells and were most likely of parasite origin. Parasite-induced differences in protein abundance concerned distinct functional categories, with most proteins being involved in host cell metabolism, cell structure, protein fate and gene transcription. Some of the modulated molecules also indicated regulatory processes on the level of host cell stress response (HSP70, HSP90), host cell apoptosis (caspase 8) and actin elongation/depolymerization (α-actinin-1, gelsonin, tropomodulin-3, transgelin). Since merozoites I were already released shortly after cell sampling, the current data reflect the situation at the end of first merogony. This is the first proteomic approach on E. bovis-infected host cells that was undertaken to gain a rather broad insight into Eimeria-induced host cell modulation. The data processed in this investigation should provide a useful basis for more detailed analyses concerning Eimeria-host cell interactions.
Taubert, Anja; Hermosilla, Carlos; Sühwold, Anke; Zahner, Horst
Cellular immune responses against Eimeria bovis are highly specific and a key factor for the development of protection against challenge infections. In this study we investigate the cellular immune responses of E. bovis primary and challenge infected calves stimulated in vitro by E. bovis merozoite I-antigen. Primary infection was accompanied by an increase of IFN-gamma and IL-2 gene transcription in whole blood samples, peaking during prepatency (8-12 days p.i.) and declining thereafter, whereas IL-4 gene transcription was induced predominantly in patency. IL-10 mRNA was not influenced by E. bovis infection. Both CD4+ and CD8+ T cells were identified as source of IFN-gamma gene transcripts, whilst IL-2 and IL-4 gene transcription was enhanced mainly in CD4+ T cells. Increased levels of IFN-gamma transcripts and protein were also found in lymphocytes isolated from ileocaecal lymph node biopsy 8 days p.i., and in cell culture supernatants obtained from antigen-stimulated peripheral blood mononuclear cells (PBMC) at days 8 and 12 p.i., respectively. Challenge infections of calves influenced neither IFN-gamma nor IL-2 gene transcription in peripheral blood or in lymph node-derived lymphocytes. In contrast, IL-4 gene transcription was increased in lymphocytes isolated from draining lymph nodes. Besides antigen-specific reactions we also found an infection-triggered induction of the non-specific activation state of PBMC in the course of primary infection as measured by the intracellular IFN-gamma and IL-4 content of phorbol-12-myristate-13-acetate/ionomycin-stimulated PBMC. This may represent a new mechanism of immune cells of E. bovis-infected calves contributing to ongoing immune reactions.
Guo, Mengyao; Wang, Guoqing; Lv, Tingting; Song, Xiaojing; Wang, Tiancheng; Xie, Guanghong; Cao, Yongguo; Zhang, Naisheng; Cao, Rongfeng
Mycoplasma bovis infection can cause endometrial inflammation leading to infertility and involuntary culling in dairy cows. Because extracellular matrix (ECM) proteins affect the adherence of mycoplasma to eukaryotic cell surface, they may play a role in the pathogenesis of the bacteria. The objective of the present study was to evaluate the endometrial inflammatory response and ECM protein expression induced by M bovis. Endometrial concentrations of inflammatory cytokines tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, and mRNA and protein expression of collagen IV (CL-IV), fibronectin (FN), and laminin (LN) were evaluated 10, 20, and 30 days after M bovis intrauterine infusion in breed cows 18 days postpartum. The presence of the bacteria in the uterus was detected by nested polymerase chain reaction and denaturing gradient gel electrophoresis. Endometrial TNF-α, IL-1β, and IL-6 concentrations in the treatment group were greater (P < 0.05) than in the positive and negative control groups 20 and 30 days after infusion. Endometrial CL-IV, FN, and LN mRNA and protein expression increased (P < 0.01) 20 days after infusion in all groups. However, the increase was more pronounced in the treatment group and reactive expressions were greater (P < 0.05) than in the positive and negative control groups 10, 20, and 30 days after infusion. In conclusion, M bovis triggered endometrial inflammatory response and increased CL-IV, FN, and LN mRNA and protein expression. The abnormal expression of ECM these proteins may promote the pathogenic effects of M bovis that lead to endometrial tissue damage and infertility.
Lerner, Uri; Amram, Eytan; Ayling, Roger D; Mikula, Inna; Gerchman, Irena; Harrus, Shimon; Teff, Dina; Yogev, David; Lysnyansky, Inna
The molecular mechanism of acquired resistance to the 16-membered macrolides tylosin (Ty) and tilmicosin (Tm) was investigated in Mycoplasma bovis field isolates. Sequence analysis of domains II and V of the two 23S rRNA alleles and ribosomal proteins L4 and L22 was performed on 54 M. bovis isolates showing different minimal inhibitory concentrations (MIC). The presence of any one of the point mutations G748A, C752T, A2058G, A2059G or A2059C (Escherichia coli numbering) in one or both alleles of the 23S rRNAs was correlated with decreased susceptibility to Ty (8-1024 μg/ml) and to Tm (32 to >256 μg/ml) in 27/27 and 27/31 M. bovis isolates, respectively. Although a single mutation in domain II or V could be sufficient to cause decreased susceptibility to Ty, our data imply that a combination of mutations in two domains is necessary to achieve higher MICs (≥ 128 μg/ml). The influence of a combination of mutations in two domains II and V on enhancement of resistance to Tm was less clear. In addition, the amino acid (aa) substitution L22-Q90H was found in 24/32 representative M. bovis isolates with different MICs, but no correlation with decreased susceptibility to Ty or Tm was identified. Multiple aa substitutions were also identified in the L4 protein, including at positions 185-186 (positions 64 and 65 in E. coli) which are adjacent to the macrolide-binding site. This is the first description of the molecular mechanism of acquired resistance to the 16-membered macrolides in M. bovis.
Wilson, D J; Justice-Allen, A; Goodell, G; Baldwin, T J; Skirpstunas, R T; Cavender, K B
The objective of this study was to evaluate the possible transmission of Mycoplasma bovis from positive sand bedding to naïve dairy calves. Twelve preweaned Holstein bull calves were blocked in pairs and randomly assigned as unexposed controls (n=6) bedded with control sand, or exposed calves (n=6) bedded with sand previously positive for M. bovis at a dairy farm. Bedding sand was cultured weekly. Nasal and ear swabs and sera were collected weekly, tracheal swabs were collected monthly, and by the end of the 105-d study, all calves were euthanized (n=10) or died (n=2). Sera were tested for M. bovis-specific antibody. Mycoplasma spp. culture was performed on nasal and ear swabs; culture and a PCR differentiating multiple Mycoplasma spp. were performed on postmortem samples of lung, retropharyngeal lymph node, and trachea from each calf. A complete necropsy also was performed. During 6 wk, mycoplasma concentration in exposed group sand was between 200 and 32,000 cfu/g. All 166 tracheal swabs, nasal and ear swabs, and postmortem tests from all calves were negative for mycoplasma. All 94 sera were negative for M. bovis-specific antibody. No gross pathology suggestive of mycoplasma disease was detected. The probability of mycoplasma detection, if an exposed calf had become infected 4 wk after exposure, ranged between 97 and 99% depending on time of exposure for individual calves. There was no evidence that sand bedding contaminated with M. bovis might serve as a source of transmission to naïve dairy calves.
Keet, D F; Michel, A L; Bengis, R G; Becker, P; van Dyk, D S; van Vuuren, M; Rutten, V P M G; Penzhorn, B L
African lions in the southern half of Kruger National Park (KNP) are infected with Mycobacterium bovis. Historically, reliable detection of mycobacteriosis in lions was limited to necropsy and microbiological analysis of lesion material collected from emaciated and ailing or repeat-offender lions. We report on a method of cervical intradermal tuberculin testing of lions and its interpretation capable of identifying natural exposure to M. bovis. Infected lions (n=52/95) were identified by detailed necropsy and mycobacterial culture. A large proportion of these confirmed infected lions (45/52) showed distinct responses to bovine tuberculin purified protein derivative (PPD) while responses to avian tuberculin PPD were variable and smaller. Confirmed uninfected lions from non-infected areas (n=11) responded variably to avian tuberculin PPD only. Various non-tuberculous mycobacteria (NTM) were cultured from 45/95 lions examined, of which 21/45 were co-infected with M. bovis. Co-infection with M. bovis and NTM did not influence skin reactions to bovine tuberculin PPD. Avian tuberculin PPD skin reactions were larger in M. bovis-infected lions compared to uninfected ones. Since NTM co-infections are likely to influence the outcome of skin testing, stricter test interpretation criteria were applied. When test data of bovine tuberculin PPD tests were considered on their own, as for a single skin test, sensitivity increased (80.8-86.5%) but false positive rate for true negatives (18.75%) remained unchanged. Finally, the adapted skin test procedure was shown not to be impeded by persistent Feline Immunodeficiency Virus(Ple) co-infection.
Mackintosh, Colin G.; Qureshi, Tariq; Waldrup, Ken; Labes, Robert E.; Dodds, Ken G.; Griffin, J. Frank T.
Tuberculosis (Tb) caused by Mycobacterium bovis is a worldwide threat to livestock and humans. One control strategy is to breed livestock that are more resistant to Mycobacterium bovis. In a 3-year heritability study 6 farmed red deer stags were selected from 39 on the basis of their differing responses to experimental challenge via the tonsillar sac with approximately 500 CFU of M. bovis. Two stags remained uninfected, two were moderately affected, and two developed serious spreading Tb. Seventy offspring, bred from these six stags by artificial insemination using stored semen, were similarly challenged with M. bovis. The offspring showed patterns of response to M. bovis challenge similar to those of their sires, providing evidence for a strong genetic basis to resistance to Tb, with an estimated heritability of 0.48 (standard error, 0.096; P < 0.01). This is the first time the heritability of Tb resistance in domestic livestock has been measured. The breeding of selection lines of resistant and susceptible deer will provide an ideal model to study the mechanisms of Tb resistance in a ruminant and could provide an additional strategy for reducing the number and severity of outbreaks of Tb in farmed deer herds. Laboratory studies to identify genetic and immunological markers for resistance to Tb are under way. Preliminary studies showed no associations between NRAMP or DRB genes and resistance to Tb in deer. Patterns of immune responses seen in resistant animals suggest that both innate and acquired pathways of immunity are necessary to produce the resistant phenotype. PMID:10678981
Cunha, Mónica V; Azorín, Beatriz; Peñuela, Rocío G; Albuquerque, Teresa; Botelho, Ana
Anthropogenic activities have cumulatively led to the dramatic decline of world populations of vultures that currently face serious survival challenges in several regions of the world. In Portugal, the three resident species qualify as endangered and are under conservation efforts, mainly in the central east and south-east regions, where habitat protection and artificial feeding stations were implemented. Concurrently, the areas under protection are highly affected by tuberculosis (TB) in cattle and wild ungulates, whose potentially infected carcasses may naturally or artificially be used as feed by local vultures. In this work, we opportunistically surveyed populations of Eurasian griffon (Gyps fulvus) and Eurasian black vulture (Aegypius monachus) for the presence of Mycobacterium bovis. Nine pathogenic mycobacteria, including one M. bovis isolate, were cultured from the oropharynx of nine of the surveyed vultures (n = 55), sampled in recovery centres or in artificial feeding stations. Genotyping of the M. bovis strain indicated spoligotype SB0121, the most frequent type in Portugal, and a unique MIRU-VNTR profile that differed in two loci from the profiles of SB0121 bovine and deer strains from the same geographical area. The M. bovis-positive griffon exhibited poor clinical condition when admitted to the recovery centre; however, clinical evidence of TB was not present. Although the significance of M. bovis isolation in this vulture specimen could not be ascertained and despite the accepted notion that vultures are naturally resistant to microbial pathogens, the sanitary follow-up of Accipitridae vulture populations in TB-hotspot areas is essential to safeguard ongoing conservation efforts and also to evaluate the suitability of standing legislation on deliberate supplementary feeding schemes for menaced birds of prey.
Pardo, J.; Carranza, C.; Turrientes, M. C.; Arellano, J. L. Pérez; Vélez, R. López; Ramajo, V.; Muro, A.
Immunodiagnostic methods based on the detection of antibodies continue to be the most effective and practical methods for the diagnosis of imported schistosomiasis. Schistosoma bovis is a species whose final natural hosts are bovines, ovines, caprines, and small wild ruminants. Different studies have demonstrated the analogies existing between S. bovis and other Schistosoma species which affect humans. The objective of this work was to evaluate the utility of S. bovis adult worm antigens (AWA) for the diagnosis of imported human schistosomiasis by enzyme-linked immunosorbent assay (ELISA) and electroimmunotransfer blotting (EITB) techniques. By detecting eggs, the ELISA for S. bovis AWA was able to definitively detect imported cases with a sensitivity of 94%. The specificity of the ELISA for S. bovis AWA was 97%. There were no differences between the results of the S. bovis AWA ELISA for patients infected with Schistosoma mansoni and those infected with Schistosoma haematobium. The EITB technique showed bands of 85, 37, and 20 kDa, which are characteristic of infections with Schistosoma spp. Specific bands to indicate infection by different species of Schistosoma have not been detected. The combined use of the ELISA for S. bovis AWA and EITB increased the global sensitivity of the study to 97%. Our findings suggest that the ELISA for S. bovis AWA is a useful test for the immunodiagnosis of imported schistosomiasis and that EITB for detecting S. bovis AWA permits the confirmation of diagnosis when the ELISA for S. bovis AWA is positive. PMID:15539523
Pardo, J; Carranza, C; Turrientes, M C; Pérez Arellano, J L; López Vélez, R; Ramajo, V; Muro, A
Immunodiagnostic methods based on the detection of antibodies continue to be the most effective and practical methods for the diagnosis of imported schistosomiasis. Schistosoma bovis is a species whose final natural hosts are bovines, ovines, caprines, and small wild ruminants. Different studies have demonstrated the analogies existing between S. bovis and other Schistosoma species which affect humans. The objective of this work was to evaluate the utility of S. bovis adult worm antigens (AWA) for the diagnosis of imported human schistosomiasis by enzyme-linked immunosorbent assay (ELISA) and electroimmunotransfer blotting (EITB) techniques. By detecting eggs, the ELISA for S. bovis AWA was able to definitively detect imported cases with a sensitivity of 94%. The specificity of the ELISA for S. bovis AWA was 97%. There were no differences between the results of the S. bovis AWA ELISA for patients infected with Schistosoma mansoni and those infected with Schistosoma haematobium. The EITB technique showed bands of 85, 37, and 20 kDa, which are characteristic of infections with Schistosoma spp. Specific bands to indicate infection by different species of Schistosoma have not been detected. The combined use of the ELISA for S. bovis AWA and EITB increased the global sensitivity of the study to 97%. Our findings suggest that the ELISA for S. bovis AWA is a useful test for the immunodiagnosis of imported schistosomiasis and that EITB for detecting S. bovis AWA permits the confirmation of diagnosis when the ELISA for S. bovis AWA is positive.
Macuamule, C L S; Wiid, I J; van Helden, P D; Tanner, M; Witthuhn, R C
Mycobacterium bovis that causes Bovine tuberculosis (BTB) can be transmitted to humans thought consumption of raw and raw fermented milk products from diseased animals. Lactic acid bacteria (LAB) used in popular traditional milk products in Africa produce anti-microbial compounds that inhibit some pathogenic and spoilage bacteria. M. bovis BCG is an attenuated non-pathogenic vaccine strain of M. bovis and the aim of the study was to determine the effect of the fermentation process on the survival of M. bovis BCG in milk. M. bovis BCG at concentrations of 6 log CFU/ml was added to products of kefir fermentation. The survival of M. bovis BCG was monitored at 12-h intervals for 72 h by enumerating viable cells on Middlebrook 7H10 agar plates enriched with 2% BD BACTEC PANTA™. M. bovis BCG was increasingly reduced in sterile kefir that was fermented for a period of 24h and longer. In the milk fermented with kefir grains, Lactobacillus paracasei subsp. paracasei or Lactobacillus casei, the viability of M. bovis BCG was reduced by 0.4 logs after 24h and by 2 logs after 48 h of fermentation. No viable M. bovis BCG was detected after 60 h of fermentation. Results from this study show that long term fermentation under certain conditions may have the potential to inactivate M. bovis BCG present in the milk. However, to ensure safety of fermented milk in Africa, fermentation should be combined with other hurdle technologies such as boiling and milk pasteurisation.
Background Mycoplasma bovis (M. bovis) is a major, but often overlooked, pathogen documented to cause respiratory disease, mastitis, and arthritis in cattle throughout China since 2008. Here, we report the development of a direct competitive enzyme-linked immunosorbent assay (Dc-ELISA) to detect M. bovis antibody. Results We used a recombinant P48 protein and monoclonal antibody (mAb) 10E. MAb 10E, prepared against the recombinant P48 protein of M. bovis, identified all M. bovis strains with no cross-reactivity with other related pathogens. Coating micro plates with P48 protein instead of whole M. bovis cells as well as the use of mAb 10E produced a specific and sensitive Dc-ELISA for M. bovis antibody detection with a cut-off percent inhibition (PI) value of 32%. Compared with two commercial indirect ELISA (i-ELISA) kits, our Dc-ELISA offered a higher positive detection rate in 165 clinical bovine serum samples. Conclusions A rapid, sensitive, and reliable serological diagnosis method was developed for M. bovis, which can facilitate M. bovis surveillance, assisting researchers in understanding the ecology and epidemiology of M. bovis. PMID:24533468
Battsetseg, Badgar; Lucero, Susana; Xuan, Xuenan; Claveria, Florencia; Byambaa, Badarch; Battur, Banzragch; Boldbaatar, Damdinsuren; Batsukh, Zayat; Khaliunaa, Tsevegmed; Battsetseg, Gonchigoo; Igarashi, Ikuo; Nagasawa, Hideyuki; Fujisaki, Kozo
Babesia equi (EMA-1) and Babesia caballi (BC48) gene fragments were amplified by polymerase chain reaction (PCR), in blood samples, and partially fed-females and egg and larval progenies of Dermacentor nuttalli, collected from horses in Altanbulag, Tuv Province, Mongolia. While Babesia parasite DNA was detected in some horse blood samples during the first PCR, all positive cases in partially fed-female ticks, eggs and larvae were confirmed by nested PCR. Present study reinforces earlier similar findings in unfed D. nuttalli ticks collected from an open space vegetation in Bayanonjuul, Tuv Province in Central Mongolia, pointing to the most likely important role of D. nuttalli in the transmission of equine babesiosis in Mongolia. The detection of parasite DNA in eggs and larval progenies is likewise suggestive of transovarial parasite transmission in this tick species.
Vega-Manriquez, X; López-Vidal, Y; Moran, J; Adams, L G; Gutiérrez-Pabello, J A
Mycobacterium tuberculosis complex species survive and replicate in phagosomes of the host cell. Cell death (CD) has been highlighted as one of the probable outcomes in this host-pathogen interaction. Previously, our group demonstrated macrophage apoptosis as a consequence of Mycobacterium bovis infection. In this study, we aimed to identify the contribution of apoptotic effector elements in M. bovis-induced CD. Bovine macrophages were either infected with M. bovis (multiplicity of infection, 10:1) or treated with an M. bovis cell extract (CFE). Structural changes compatible with CD were evaluated. Chromatin condensation was increased three times by the CFE. On the other hand, a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assay demonstrated that levels of DNA fragmentation induced by M. bovis and CFE were 53.7% +/- 24% and 38.9% +/- 14%, respectively, whereas control cells had a basal proportion of 8.9% +/- 4.1%. Rates of DNA fragmentation were unaffected by the presence of the pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp (z-VAD). Cells treated with 100 mug of CFE for 12 h had a fivefold decrease in the level of mitochondrial outer membrane permeabilization compared to that of untreated cells. Neither M. bovis infection nor CFE treatment induced activation of caspase 3, 8, or 9. Translocation of apoptosis-inducing factor (AIF) to the nucleus was identified in 32% +/- 3.5% and 26.3% +/- 4.9% of M. bovis-infected and CFE-treated cells, respectively. Incubation of macrophages with z-VAD prior to infection did not alter the percentage of cells showing AIF translocation. Our data suggest that M. bovis-induced CD in bovine macrophages is caspase independent with AIF participation.
Cui, Yongyong; Zhao, Deming; Sreevatsan, Srinand; Liu, Chunfa; Yang, Wei; Song, Zhiqi; Yang, Lifeng; Barrow, Paul; Zhou, Xiangmei
Mycobacterium bovis (M. bovis) is highly adapted to macrophages and has developed multiple mechanisms to resist intracellular assaults. However, the host cells in turn deploy a multipronged defense mechanism to control bacterial infection. Endoplasmic reticulum (ER) stress-mediated apoptosis is one such primary defense mechanism. However, the role of interferon regulatory factor 3 (IRF3) between ER stress and apoptosis during M. bovis infection is unknown. Here, we demonstrate that M. bovis effectively induced apoptosis in murine macrophages. Caspase-12, caspase-9, and caspase-3 were activated over a 48 h infection period. The splicing of XBP-1 mRNA and the level of phosphorylation of eIF2α, indicators of ER stress, significantly increased at early time points after M. bovis infection. The expansion of the ER compartment, a morphological hallmark of ER stress, was observed at 6 h. Pre-treatment of Raw 264.7 cells with 4-PBA (an ER stress-inhibitor) reduced the activation of the ER stress indicators, caspase activation and its downstream poly (ADP-ribose) polymerase (PARP) cleavage, phosphorylation of TBK1 and IRF3 and cytoplasmic co-localization of STING and TBK1. M. bovis infection led to the interaction of activated IRF3 and cytoplasmic Bax leading to mitochondrial damage. Role of IRF3 in apoptosis was further confirmed by blocking this molecule with BX-795 that showed significant reduction expression of caspase-8 and caspase-3. Intracellular survival of M. bovis increased in response to 4-PBA and BX-795. These findings indicate that STING-TBK1-IRF3 pathway mediates a crosstalk between ER stress and apoptosis during M. bovis infection, which can effectively control intracellular bacteria. PMID:28018864
Badawy, Ahmed Ibrahem I; Lutz, Kathleen; Taubert, Anja; Zahner, Horst; Hermosilla, Carlos
Host immune responses conducted against antigens of Eimeria bovis are key factors for the development of protective immunity against this protozoan disease. In this study we investigated the expression of E. bovis-derived antigens on the host cell surface membrane during E. bovis first merogony in vitro. Host cells carrying E. bovis-meront I stages expressed E. bovis host cell surface antigens (EbHCSAg) on their surface membrane which were recognised by hyperimmune sera of calves and by sera from rats immunized with E. bovis merozoites I, when tested by indirect immune fluorescent antibody test (IIFAT), laser scanning confocal microscopy (LSCM) and immune electron microscopy. Expression of EbHCSAg on permissive host cells was earliest detected 7 days p. i., thus coinciding with the onset of the parasite replication. Membrane-associated EbHCSAg were removed from infected host cells by proteinase K, partially by Triton X-100, Triton X-114 and Triton X-405, but not by 1 M NaCl, CHAPS or phospholipase C treatment. Antibodies, affinity-purified on paraformaldehyde/glutardialdehyde (PAGA)-fixed E. bovis meront I-infected bovine host cells bound to the surface meront I-carrying cells and to merozoites I (IIFAT, LSCM) but, in contrast to untreated sera, not to sporozoites. When tested on methanol-fixed merozoites I and sporozoites by IIFAT, affinity-purified antibodies bound to structures in the apical complex area of merozoites I, but not to sporozoites, whilst untreated sera caused diffuse labelling of internal structures of both parasite stages. Immune electron microscopy demonstrated binding of affinity-purified antibodies to micronemes and dense granules of merozoites I. Although the function of EbHCSAg is still unknown, results of this study might suggest an involvement in the development of protective immunity against E. bovis infections.
Roug, Annette; Geoghegan, Claire; Wellington, Elizabeth; Miller, Woutrina A; Travis, Emma; Porter, David; Cooper, David; Clifford, Deana L; Mazet, Jonna A K; Parsons, Sven
A real-time PCR protocol for detecting Mycobacterium bovis in feces was evaluated in bovine tuberculosis-infected African buffalo (Syncerus caffer). Fecal samples spiked with 1.42 × 10(3) cells of M. bovis culture/g and Bacille Calmette-Guérin standards with 1.58 × 10(1) genome copies/well were positive by real-time PCR but all field samples were negative.
Stewart, Linda D.; McNair, James; McCallan, Lyanne; Gordon, Alan; Grant, Irene R.
Immunomagnetic separation (IMS) can selectively isolate and concentrate Mycobacterium bovis cells from lymph node tissue to facilitate subsequent detection by PCR (IMS-PCR) or culture (IMS-MGIT). This study describes application of these novel IMS-based methods to test for M. bovis in a survey of 280 bovine lymph nodes (206 visibly lesioned (VL), 74 non-visibly lesioned (NVL)) collected at slaughter as part of the Northern Ireland bovine TB eradication programme. Their performance was evaluated relative to culture. Overall, 174 (62.1%) lymph node samples tested positive by culture, 162 (57.8%) by IMS-PCR (targeting IS6110), and 191 (68.2%) by IMS-MGIT culture. Twelve (6.9%) of the 174 culture positive lymph node samples were not detected by either of the IMS-based methods. However, an additional 79 M. bovis positive lymph node samples (27 (13.1%) VL and 52 (70.3%) NVL) were detected by the IMS-based methods and not by culture. When low numbers of viable M. bovis are present in lymph nodes (e.g. in NVLs of skin test reactor cattle) decontamination prior to culture may adversely affect viability, leading to false negative culture results. In contrast, IMS specifically captures whole M. bovis cells (live, dead or potentially dormant) which are not subject to any deleterious treatment before detection by PCR or MGIT culture. During this study only 2.7% of NVL lymph nodes tested culture positive, whereas 70.3% of the same samples tested M. bovis positive by the IMS-based tests. Results clearly demonstrate that not only are the IMS-based methods more rapid but they have greater detection sensitivity than the culture approach currently used for the detection of M. bovis infection in cattle. Adoption of the IMS-based methods for lymph node testing would have the potential to improve M. bovis detection in clinical samples. PMID:23469275
Kupferwasser, I; Darius, H; Muller, A; Mohr-Kahaly, S; Westermeier, T; Oelert, H; Erbel, R; Meyer, J
Aim—To compare the clinical and morphological characteristics of patients with Streptococcus bovis endocarditis with those of patients with endocarditis caused by other microorganisms. Methods—177 consecutive patients (Streptococcus bovis, 22; other streptococci, 94; staphylococci, 44; other, 17) with definite infective endocarditis according to the Duke criteria were included. All patients underwent transthoracic and transoesophageal echocardiography. In 88 patients, findings from surgery/necropsy were obtained. Results—S bovis endocarditis was associated with older patients, with a higher mortality (p = 0.04), and with a higher rate of cardiac surgery (p < 0.001) than other microorganisms, although embolic events were observed less often (p = 0.02). Pathological gastrointestinal lesions were detected in 45% of the patients. Multiple valves were affected in 68% of the patients with S bovis endocarditis and in 20% of those with other organisms (p < 0.001). Moderate or severe regurgitation occurred more often in S bovis endocarditis than with other microorganisms (p = 0.05). When surgery or necropsy was performed, infectious myocardial infiltration of the left ventricle was confirmed histopathologically in 36% of the patients with S bovis endocarditis and in 10% of those with other organisms (p = 0.002). Conclusions—S bovis endocarditis is a severe illness because of the more common involvement of multiple valves, and of the frequent occurrence of haemodynamically relevant valvar regurgitation and infectious myocardial infiltration. Keywords: infective endocarditis; Streptococcus bovis; transoesophageal echocardiography; valvar disease PMID:9875088
Becker, Claire A M; Thibault, François M; Arcangioli, Marie-Anne; Tardy, Florence
Mycoplasma (M.) bovis has recently emerged as a major, worldwide etiological agent of bovine respiratory diseases leading to huge economic losses mainly due to high morbidity and mortality as well as poor growth rates. The spread of M. bovis infections between different animals, herds, regions or countries has been often reported to be connected to the movement of animals. However, despite recent considerable efforts, no universal subtyping method is yet available to trace M. bovis isolates circulation at an international scale. Moreover in France, the overall population diversity of M. bovis isolates has not been assessed since the early 1990s. This study was conducted to fill in these gaps. The genotypic diversity between sixty isolates collected in France over the last 35 years was assessed using two molecular subtyping methods that addressed either the long-term epidemiological relationships (Multi Locus Sequence Typing, MLST) or the genetic microvariations (Multiple Locus VNTR Analysis, MLVA) between isolates. Phenotypic diversity was also analyzed by using Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) to compare the main protein patterns of isolates. All proposed subtyping approaches were optimized and led to the same pattern in the French M. bovis population that consisted of two clusters, the first one comprising isolates collected before 2000 and the second, those collected after 2000. Recent strains were further shown to be more homogeneous than older ones, which is consistent with the spread of a single clone throughout the country. Because this spread was concomitant with the emergence of multiresistant M. bovis isolates, several hypotheses are discussed to explain the homogeneity of M. bovis isolates in France, even though the M. bovis species is fully equipped to generate diversity.
Seim, O.S.; Filewicz, E.C.; Hutter, E.
An irradiation subassembly for use in a nuclear reactor is described which includes a bundle of slender elongated irradiation -capsules or fuel elements enclosed by a coolant tube and having yieldable retaining liner between the irradiation capsules and the coolant tube. For a hexagonal bundle surrounded by a hexagonal tube the yieldable retaining liner may consist either of six segments corresponding to the six sides of the tube or three angular segments each corresponding in two adjacent sides of the tube. The sides of adjacent segments abut and are so cut that metal-tometal contact is retained when the volume enclosed by the retaining liner is varied and Springs are provided for urging the segments toward the center of the tube to hold the capsules in a closely packed configuration. (Official Gazette)
Zanine, Anderson de Moura; Bonelli, Emerson Alencar; de Souza, Alexandre Lima; Ferreira, Daniele de Jesus; Santos, Edson Mauro; Ribeiro, Marinaldo Divino; Geron, Luiz Juliano Valério; Pinho, Ricardo Martins Araujo
This study aimed to evaluate the effects of Streptococcus bovis on the fermentation characteristics and nutritive value of Tanzania grass silage. Tanzania grass was chopped and left untreated (U) or treated with Streptococcus bovis JB1 at 1 × 106 colony-forming units per gram (cfu/g) of fresh forage or Streptococcus bovis HC5 at 1 × 106 cfu/g of fresh forage and packed into sixtuplicate laboratory silos. The largest number of enterobacteria, molds and yeast (M&Y) occurred in untreated silages and the smallest populations of enterobacteria and M&Y and the largest numbers of lactic acid bacteria (LAB), at 9.81 and 9.87 log cfu/g, were observed in Streptococcus bovis JB1 and HC5, respectively (P < 0.05). Silages treated with JB1 and HC5 had lower (P < 0.05) silage pHs and concentrations of ammoniacal nitrogen (NH3-N) than untreated silages. The application of Streptococcus bovis JB1 and HC5 resulted in fewer losses through gases and effluents (P < 0.05), which resulted in greater dry matter recovery (DMR) and crude protein recovery (CPR) (P < 0.05). Streptococcus bovis JB1 and HC5 improved the fermentative profile and increased the concentration of crude protein and DMR and CPR in Tanzania grass silage. PMID:27073806
Su, Feng; Wang, Yongsheng; Liu, Guanghui; Ru, Kun; Liu, Xin; Yu, Yuan; Liu, Jun; Wu, Yongyan; Quan, Fusheng; Guo, Zekun; Zhang, Yong
Bovine tuberculosis results from infection with Mycobacterium bovis, a member of the Mycobacterium tuberculosis family. Worldwide, M. bovis infections result in economic losses in the livestock industry; cattle production is especially hard-hit by this disease. Generating M. bovis-resistant cattle may potentially mitigate the impact of this disease by reducing M. bovis infections. In this study, we used transgenic somatic cell nuclear transfer to generate cattle expressing the gene encoding human β-defensin 3 (HBD3), which confers resistance to mycobacteria in vitro. We first generated alveolar epithelial cells expressing HBD3 under the control of the bovine MUC1 promoter, and confirmed that these cells secreted HBD3 and possessed anti-mycobacterial capacity. We then generated and identified transgenic cattle by somatic cell nuclear transfer. The cleavage and blastocyst formation rates of genetically modified embryos provided evidence that monoclonal transgenic bovine fetal fibroblast cells have an integral reprogramming ability that is similar to that of normal cells. Five genetically modified cows were generated, and their anti-mycobacterial capacities were evaluated. Alveolar epithelial cells and macrophages from these cattle expressed higher levels of HBD3 protein compared with non-transgenic cells and possessed effective anti-mycobacterial capacity. These results suggest that the overall risk of M. bovis infection in transgenic cattle is efficiently reduced, and support the development of genetically modified animals as an effective tool to reduce M. bovis infection.
Fenton, Karla A; Fitzgerald, Scott D; Bolin, Steve; Kaneene, John; Sikarskie, James; Greenwald, Rena; Lyashchenko, Konstantin
An endemic focus of Mycobacterium bovis (M. bovis) infection in the state of Michigan has contributed to a regional persistence in the animal population. The objective of this study was to determine if Virginia opossums (Didelphis virginiana) contribute to disease persistence by experimentally assessing intraspecies lateral transmission. One wild caught pregnant female opossum bearing 11 joeys (young opossum) and one age-matched joey were obtained for the study. Four joeys were aerosol inoculated with M. bovis (inoculated), four joeys were noninoculated (exposed), and four joeys plus the dam were controls. Four replicate groups of one inoculated and one exposed joey were housed together for 45 days commencing 7 days after experimental inoculation. At day 84 opossums were sacrificed. All four inoculated opossums had a positive test band via rapid test, culture positive, and gross/histologic lesions consistent with caseogranulomatous pneumonia. The exposed and control groups were unremarkable on gross, histology, rapid test, and culture. In conclusion, M. bovis infection within the inoculated opossums was confirmed by gross pathology, histopathology, bacterial culture, and antibody tests. However, M. bovis was not detected in the control and exposed opossums. There was no appreciable lateral transmission of M. bovis after aerosol inoculation and 45 days of cohabitation between infected and uninfected opossums.
Sylvester, Tashnica Taime; Martin, Laura Elizabeth Rosen; Buss, Peter; Loxton, Andre Gareth; Hausler, Guy Anton; Rossouw, Leana; van Helden, Paul; Parsons, Sven David Charles; Olea-Popelka, Francisco; Miller, Michele Ann
Mycobacterium bovis, the causative agent of bovine tuberculosis (BTB), is endemic in the Kruger National Park (KNP), South Africa. African lions ( Panthera leo ) are susceptible to BTB, but the impact of the disease on lion populations is unknown. In this study, we used a novel gene expression assay for chemokine (C-X-C motif) ligand 9 (CXCL9) to measure the prevalence of M. bovis infection in 70 free-ranging lions that were opportunistically sampled in the southern and central regions of the KNP. In the southern region of the KNP, the apparent prevalence of M. bovis infection was 54% (95% confidence interval [CI]=36.9-70.5%), compared with 33% (95% CI=18.0-51.8%) in the central region, an important difference (P=0.08). Prevalence of M. bovis infection in lions showed similar patterns to estimated BTB prevalence in African buffaloes ( Syncerus caffer ) in the same areas. Investigation of other risk factors showed a trend for older lions, males, or lions with concurrent feline immunodeficiency virus infection to have a higher M. bovis prevalence. Our findings demonstrate that the CXCL9 gene expression assay is a useful tool for the determination of M. bovis status in free-ranging lions and identifies important epidemiologic trends for future studies.
Jang, Hyeung-Jin; Nde, Chantal; Toghrol, Freshteh; Bentley, William E
Mycobacterium bovis BCG strain Pasteur 1173P2 responds with adaptive and protective strategies against oxidative stress. Despite advances in our understanding of the responses to oxidative stress in many specific cases, the connectivity between targeted protective genes and the rest of cell metabolism remains obscure. This study was therefore carried out to investigate the genome-wide response of M. bovis BCG to hydrogen peroxide after 10 and 60 min of treatment. ATP measurements were carried out in order to monitor the changes in M. bovis BCG growth over a 1 h period. The furA gene in Mycobacterium bovis, a pleiotropic regulator that couples iron metabolism to the oxidative stress response was involved in the response to hydrogen peroxide stress. There were also increased levels of catalase/ peroxidase (KatG) and the biosynthesis operon of mycobactin. This study revealed significant upregulation of the oxidative response group of M. bovis, amino acid transport and metabolism, defense mechanisms, DNA replication, recombination and repair, and downregulation of cell cycle control, mitosis, and meiosis, lipid transport and metabolism, and cell wall/membrane biogenesis. This study shows that the treatment of M. bovis BCG with hydrogen peroxide induces iron acquisition related genes and oxidative stress response genes within one hour of treatment.
Vázquez, Cristina Lourdes; Bianco, María Verónica; Blanco, Federico Carlos; Forrellad, Marina Andrea; Gutierrez, Maximiliano Gabriel
ABSTRACT Mycobacterium bovis causes tuberculosis in a wide variety of mammals, with strong tropism for cattle and eventually humans. P27, also called LprG, is among the proteins involved in the mechanisms of the virulence and persistence of M. bovis and Mycobacterium tuberculosis. Here, we describe a novel function of P27 in the interaction of M. bovis with its natural host cell, the bovine macrophage. We found that a deletion in the p27-p55 operon impairs the replication of M. bovis in bovine macrophages. Importantly, we show for the first time that M. bovis arrests phagosome maturation in a process that depends on P27. This effect is P27 specific since complementation with wild-type p27 but not p55 fully restored the wild-type phenotype of the mutant strain; this indicates that P55 plays no important role during the early events of M. bovis infection. In addition, we also showed that the presence of P27 from M. smegmatis decreases the association of LAMP-3 with bead phagosomes, indicating that P27 itself blocks phagosome-lysosome fusion by modulating the traffic machinery in the cell host. PMID:28031264
Fenton, Karla A.; Fitzgerald, Scott D.; Bolin, Steve; Kaneene, John; Sikarskie, James; Greenwald, Rena; Lyashchenko, Konstantin
An endemic focus of Mycobacterium bovis (M. bovis) infection in the state of Michigan has contributed to a regional persistence in the animal population. The objective of this study was to determine if Virginia opossums (Didelphis virginiana) contribute to disease persistence by experimentally assessing intraspecies lateral transmission. One wild caught pregnant female opossum bearing 11 joeys (young opossum) and one age-matched joey were obtained for the study. Four joeys were aerosol inoculated with M. bovis (inoculated), four joeys were noninoculated (exposed), and four joeys plus the dam were controls. Four replicate groups of one inoculated and one exposed joey were housed together for 45 days commencing 7 days after experimental inoculation. At day 84 opossums were sacrificed. All four inoculated opossums had a positive test band via rapid test, culture positive, and gross/histologic lesions consistent with caseogranulomatous pneumonia. The exposed and control groups were unremarkable on gross, histology, rapid test, and culture. In conclusion, M. bovis infection within the inoculated opossums was confirmed by gross pathology, histopathology, bacterial culture, and antibody tests. However, M. bovis was not detected in the control and exposed opossums. There was no appreciable lateral transmission of M. bovis after aerosol inoculation and 45 days of cohabitation between infected and uninfected opossums. PMID:22701815
Bangoura, Berit; Mundt, Hans-Christian; Schmäschke, Ronald; Westphal, Bernhard; Daugschies, Arwid
The present study was designed to investigate the prevalence of the pathogenic coccidia species E. bovis and E. zuernii in shed-reared animals in German dairy and fattening facilities.Samples were obtained from 65 cattle farms distributed randomly across all the regions of Germany, regardless of the occurrence of clinical problems. The samples were obtained rectally. Faecal consistency and the total number of oocysts per gram of faeces (OPG) were determined for Eimeria spp., along with the separate OPG values for Eimeria (E.) bovis and E. zuernii. A questionnaire was completed for each farm to record information about herd size and management together with individual animal data. Eimeria oocysts, regardless of the kind of Eimeria spp., were detected in 62 of these farms, which gives a prevalence of 95.4 %. The farm prevalence of the pathogenic species was 76.9 % for E. bovis and 83.1 % for E. zuernii. The average oocyst excretion level was 2,950 OPG in terms of total Eimeria spp. oocyst excretion, 700 OPG for E. bovis and 1,500 OPG for E. zuernii.The number of oocysts excreted could not be correlated significantly with farm type or farm management but depended on the floor type which influences the infection pressure, on the age of the calves and the time after rehousing. In general, higher oocyst excretion rates were found in calves kept on litter compared to rearing on slatted floor. Younger calves and calves sampled early after housing shed higher amounts of oocysts than older calves and calves stabled a longer period before sampling, respectively. Furthermore, there was a positive correlation between OPG and the observation of diarrhoea, defined as observation of a loose to liquid faecal consistency. Excretion of E. zuernii oocysts was more closely linked to the occurrence of diarrhoea than E. bovis oocyst excretion. This study confirms that the pathogenic coccidia E. bovis and E. zuernii are ubiquitous in German cattle populations and a significant cause of
Kallio, Eva R; Begon, Michael; Birtles, Richard J; Bown, Kevin J; Koskela, Esa; Mappes, Tapio; Watts, Phillip C
Tick-borne diseases pose an increasingly important public health problem in Europe. Rodents are the reservoir host for many tick-transmitted pathogens, including Anaplasma phagocytophilum and Babesia microti, which can cause human granulocytic anaplasmosis and babesiosis, respectively. To estimate the presence of these pathogens in rodents in Finland, we examined blood samples from 151 bank voles (Myodes glareolus) and demonstrate, for the first time, that A. phagocytophilum and B. microti commonly infect bank voles (in 22% and 40% of animals, respectively) in Finland. Sequence analysis of a fragment of 18S rRNA showed that the B. microti strain isolated was identical to the Munich strain, which is considered to be nonzoonotic. The A. phagocytophilum strain (based on a fragment of the msp4 gene) was identical to one found earlier in rodents in the United Kingdom that is transmitted by the tick Ixodes trianguliceps, all the life stages of which feed on small mammals. The infection probability of B. microti in the bank voles was the greater the older the individual was, and males were more often infected than females. A. phagocytophilum infection probability first increased and then decreased with the age of individual without any difference between sexes. While these pathogens presumably pose a limited zoonotic risk to humans in Finland, they might have important interactions with other rodent pathogens and therefore affect infection dynamics of, for example, zoonotic pathogens.
Knowles, D P; Kappmeyer, L S; Perryman, L E
Horses possessing a normal immune system and spleen often control infection caused by Babesia equi. However, splenectomized horses are unable to control B. equi infection and usually succumb to the infection. To investigate the role of the spleen in the control of B. equi infection in the absence of specific immune responses, two 1-month-old foals with severe combined immunodeficiency (SCID) and two age-matched normal foals were inoculated with B. equi. The SCID foals became febrile seven days postinoculation and developed terminal parasitemias of 41 and 29%. The SCID foals had greater than 50% decreases in indices of total erythrocytes, packed-cell volumes, and hemoglobin concentrations. Both SCID foals were euthanized in extremis at 10 days postinoculation. As expected, the serum of the SCID foals lacked detectable antibodies to B. equi antigens. In contrast, the normal foals inoculated with B. equi produced detectable anti-erythrocyte-stage parasite antibodies by 7 days and controlled clinical disease by 12 days postinoculation. Although SCID foals lack functional T and B lymphocytes, they do possess complement, macrophages, granulocytes, and natural killer cells, as well as a spleen. Therefore, the data indicate that specific immune responses are required to control B. equi parasitemia but are not required for erythrocyte lysis in infected horses. Furthermore, the spleen is not able to control B. equi parasitemia in the absence of specific immune responses to parasite antigens. Images PMID:8168957
Zintl, Annetta; Mulcahy, Grace; Skerrett, Helen E.; Taylor, Stuart M.; Gray, Jeremy S.
Babesia divergens is an intraerythrocytic protozoan parasite, transmitted by the tick Ixodes ricinus, and is the main agent of bovine babesiosis in Europe. It is not only a cause of significant loss to the cattle industry; it can also infect immunocompromised humans, causing medical emergencies characterized by rapid fulmination and parasitemias that may exceed 70%. The current emphasis in Europe on sustainable agriculture and extensification is likely to lead to an increase in vector tick populations with increased risk of infection. Despite the veterinary and zoonotic importance of this parasite, relatively little research has been carried out on B. divergens, and many questions regarding the parasite's epidemiology and the host's response remain unanswered. A better understanding of the species' biology and host-parasite interactions may lead to improved control mechanisms and new trends in vaccine and antibabesial drug development. This review provides the first comprehensive summary of B. divergens biology, including its morphology, life cycle, and host specificity, and the current state of knowledge of both human and bovine infections. PMID:14557289
Zygner, Wojciech; Gójska-Zygner, Olga; Wesołowska, Agnieszka; Wędrychowicz, Halina
Urinary creatinine to serum creatinine (UCr/SCr) ratio and renal failure index (RFI) are useful indices of renal damage. Both UCr/SCr ratio and RFI are used in differentiation between prerenal azotaemia and acute tubular necrosis. In this work the authors calculated the UCr/SCr ratio and RFI in dogs infected with Babesia canis and the values of these indices in azotaemic dogs infected with the parasite. The results of this study showed significantly lower UCr/SCr ratio in dogs infected with B. canis than in healthy dogs. Moreover, in azotaemic dogs infected with B. canis the UCr/SCr ratio was significantly lower and the RFI was significantly higher than in non-azotaemic dogs infected with B. canis. The calculated correlation between RFI and duration of the disease before diagnosis and treatment was high, positive and statistically significant (r = 0.89, p < 0.001). The results of this study showed that during the course of canine babesiosis caused by B. canis in Poland acute tubular necrosis may develop.
Garg, Aprajita; Stein, Anna; Zhao, William; Dwivedi, Ankit; Frutos, Roger; Cornillot, Emmanuel; Ben Mamoun, Choukri
The apicomplexan intraerythrocytic parasite Babesia microti is an emerging human pathogen and the primary cause of human babesiosis, a malaria-like illness endemic in the United States. The pathogen is transmitted to humans by the tick vector, Ixodes scapularis, and by transfusion of blood from asymptomatic B. microti-infected donors. Whereas the nuclear and mitochondrial genomes of this parasite have been sequenced, assembled and annotated, its apicoplast genome remained incomplete, mainly due to its low representation and high A+T content. Here we report the complete sequence and annotation of the apicoplast genome of the B. microti R1 isolate. The genome consists of a 28.7 kb circular molecule encoding primarily functions important for maintenance of the apicoplast DNA, transcription, translation and maturation of organellar proteins. Genome analysis and annotation revealed a unique gene structure and organization of the B. microti apicoplast genome and suggest that all metabolic and non-housekeeping functions in this organelle are nuclear-encoded. B. microti apicoplast functions are significantly different from those of the host, suggesting that they might be useful as targets for development of potent and safe therapies for the treatment of human babesiosis.
de Rezende, Jania; Rangel, Charles P; McIntosh, Douglas; Silveira, Júlia A G; Cunha, Nathalie C; Ramos, Carlos A N; Fonseca, Adivaldo H
Cultures of tick hemocytes represent alternative cell lines for the isolation and cultivation of a variety of hemoparasites. The present study reports the development and evaluation of methods for the in vitro culture and maintenance of sporokinetes of Babesia bigemina in association with hemocytes of the tick Rhipicephalus microplus. Hemolymph, from engorged females infected with B. bigemina sporokinetes, was incubated at 28 °C in L15 culture medium supplemented with 40% fetal bovine serum. Adherence of hemocytes to flask surfaces and the development of B. bigemina sporokinetes commenced on the first day of cultivation. The protozoa demonstrated clear motility and the capacity to adhere to hemocyte membranes for up to 25 days, at which time the hemocytes began to show signs of degeneration. Examination of Giemsa stained hemocyte cultures, revealed the presence of pyriformis forms, as well as mature and immature sporokinetes with dark red nuclei, centralized or near the apical extremities. Sporokinetes harvested from culture supernatants were cryopreserved in liquid nitrogen. Inoculation of parasite-free hemocyte cultures with defrosted sporokinetes, demonstrated the viability and interaction of the protozoa with the hemocytes over 21 days. Cultured hemocytes of R. microplus hold potential for development as a tool in the study of host parasite interactions and as a substrate for the in vitro maintenance of B. bigemina sporokinetes.
Begon, Michael; Birtles, Richard J.; Bown, Kevin J.; Koskela, Esa; Mappes, Tapio; Watts, Phillip C.
Abstract Tick-borne diseases pose an increasingly important public health problem in Europe. Rodents are the reservoir host for many tick-transmitted pathogens, including Anaplasma phagocytophilum and Babesia microti, which can cause human granulocytic anaplasmosis and babesiosis, respectively. To estimate the presence of these pathogens in rodents in Finland, we examined blood samples from 151 bank voles (Myodes glareolus) and demonstrate, for the first time, that A. phagocytophilum and B. microti commonly infect bank voles (in 22% and 40% of animals, respectively) in Finland. Sequence analysis of a fragment of 18S rRNA showed that the B. microti strain isolated was identical to the Munich strain, which is considered to be nonzoonotic. The A. phagocytophilum strain (based on a fragment of the msp4 gene) was identical to one found earlier in rodents in the United Kingdom that is transmitted by the tick Ixodes trianguliceps, all the life stages of which feed on small mammals. The infection probability of B. microti in the bank voles was the greater the older the individual was, and males were more often infected than females. A. phagocytophilum infection probability first increased and then decreased with the age of individual without any difference between sexes. While these pathogens presumably pose a limited zoonotic risk to humans in Finland, they might have important interactions with other rodent pathogens and therefore affect infection dynamics of, for example, zoonotic pathogens. PMID:24848684
Guven, Esin; Avcioglu, Hamza; Deniz, Ahmet; Balkaya, İbrahim; Abay, Ugur; Yavuz, Şevki; Akyüz, Muzaffer
Equine piroplasmosis (EP) is a hemoprotozoan tick-borne disease with worldwide distribution that is caused by Theileria equi and Babesia caballi. There are studies reporting the presence of equine piroplasmosis in Turkey but the situation in Erzurum is unknown. The aim of the current study was to determine the situation of equine piroplasmosis in jeered horses in Erzurum. Between April and August 2015, a total of 125 Arabian horse were examined and blood samples were collected. At the time of sampling, animals were also examined for tick infestations and clinical signs. Besides microscopic examination of Giemsastained blood smears, multiplex PCR performed with species specific primers partially amplifying the 18S rRNA gene of B. caballi and T. equi. During the microscopic examination of blood smears, T. equi piroplasms were found in 6 (4.8%) samples. In total, 11 (8.8%) T. equi DNA were detected with multiplex PCR. B. caballi piroplasms or DNA were not obtained. BLAST analysis of the sequenced T. equi samples (GenBank: KU921661-KU921667) indicated 98.8-100% identity to each other, and 100% similarity to T. equi isolates in South Africa, Iran, China, Sudan, India, Mongolia, Trinidad, Kenya, Spain, Serbia, Bosnia and Herzegovina and Turkey (Bursa). The results of our study indicate that T. equi occurs more frequently than B. caballi in the study area. To the authors' knowledge, this is the first report of the molecular detection of equine piroplasmosis in jeered horses in Erzurum, Turkey.
Background The incidence of bovine babesiosis, caused by Babesia divergens (Apicomplexa: Piroplasmida) has decreased markedly since the 1930 s, but may re-emerge as a consequence of climate change and changes in legislation and pasturing practices. This is a potentially serious disease, with both economical and animal welfare consequences. Therefore, there is a need to survey the distribution of B. divergens. Methods We tested sera from 306 healthy pastured cows from 24 farms along the southern Norwegian coast by using an indirect immunofluorescence IgG antibody test (IFAT). Fractions of seropositive cows were compared by calculating 95% CI. Results The results of this test showed that 27% of the sera were positive for B. divergens antibodies. The fraction of antibody-positive sera that we detected showed a two-humped distribution, with a high fraction of positives being found in municipalities in the western and eastern parts of the study area, while the municipalities between these areas had few or no positive serum samples. Conclusions Neither the farmers' observations nor the Norwegian Dairy Herd Recording System give an adequate picture of the distribution of bovine babesiosis. Serological testing of cows by using IFAT is a convenient way of screening for the presence of B. divergens in an area. PMID:20925923
Zamoto-Niikura, Aya; Morikawa, Shigeru; Hanaki, Ken-Ichi; Holman, Patricia J; Ishihara, Chiaki
The U.S. lineage, one of the major clades in the Babesia microti group, is known as a causal agent of human babesiosis mostly in the northeastern and upper midwestern United States. This lineage, however, also is distributed throughout the temperate zone of Eurasia with several reported human cases, although convincing evidence of the identity of the specific vector(s) in this area is lacking. Here, the goal was to demonstrate the presence of infectious parasites directly in salivary glands of Ixodes persulcatus, from which U.S. lineage genetic sequences have been detected in Asia, and to molecularly characterize the isolates. Five PCR-positive specimens were individually inoculated into hamsters, resulting in infections in four; consequently, four strains were newly established. Molecular characterization, including 18S rRNA, β-tubulin, and CCT7 gene sequences, as well as Western blot analysis and indirect fluorescent antibody assay, revealed that all four strains were identical to each other and to the U.S. lineage strains isolated from rodents captured in Japan. The 18S rRNA gene sequence from the isolates was identical to those from I. persulcatus in Russia and China, but the genetic and antigenic profiles of the Japanese parasites differ from those in the United States and Europe. Together with previous epidemiological and transmission studies, we conclude that I. persulcatus is likely the principal vector for the B. microti U.S. lineage in Japan and presumably in northeastern Eurasia.
Machado, R Z; Toledo, C Z P; Teixeira, M C A; André, M R; Freschi, C R; Sampaio, P H
Piroplasmosis in donkeys has been recognized as a serious problem of major economic importance, since the affected animals manifest loss of appetite and decreased working capacity. The present work is aimed at detecting infection or exposure of donkeys in São Paulo, Brazil to Theileria (T.) equi and Babesia (B.) caballi using molecular and serological approaches. EDTA-blood and serum samples were collected from 88 donkeys (Equus asinus). From 88 sampled donkeys, 65 (73.86%; 95% confidence interval, PI=63.41, 82.65) and 82 (93.2%; 95% confidence interval, PI=85.75, 97.46) animals showed IgG antibodies to T. equi (by ELISA) and B. caballi (by IFAT), respectively. Twenty-eight (31.81%; 95% confidence interval, PI=22.3, 42.61) and 18 (20.45%; 95% confidence interval, PI=12.6, 30.39) donkeys were positive to T. equi and B. caballi nested PCR assays, respectively. The results indicated that T. equi and B. caballi are prevalent among donkeys in Brazil.
Davitkov, Darko; Vucicevic, Milos; Stevanovic, Jevrosima; Krstic, Vanja; Slijepcevic, Dajana; Glavinic, Uros; Stanimirovic, Zoran
Equine piroplasmosis is significant tick-borne disease with wide distribution. The prevalence of equine piroplasmosis in Serbia, Montenegro and Bosnia and Herzegovina is unknown. In aim to obtain a first insight into the prevalence we performed molecular epidemiological study which included 142 horses, on seven locations in these three countries. We first performed PCR for the detection of a 450bp long section of the 18S rRNA of piroplasma-specific region. For all positive samples we have done multiplex PCR for the species detection. Species determination was further confirmed by sequencing PCR products of 10 randomly selected Theileria equi and all Babesia caballi samples. The overall prevalence rates in analysed region for T. equi and B. caballi were 22.5% and 2.1%, respectively. Possible risk factors (such as location, age, sex and activity) associated with PCR positivity were evaluated. Marked differences were found in prevalence between geographic areas. There was no significant association between positivity and age group. T. equi was more prevalent in females and farming horses. This is the first report on the molecular survey of T. equi and B. caballi in central Balkan. Further prevalence studies on definitive host and vectors in this region are necessary.
Liu, Yonggang; Tan, Peng; Liu, Shanshan; Shi, Hang; Feng, Xin; Ma, Qun
Objective: Calculus bovis have been widely used in Chinese herbology for the treatment of hyperpyrexia, convulsions, and epilepsy. Nowadays, due to the limited source and high market price, the substitutes, artificial and in vitro cultured Calculus bovis, are getting more and more commonly used. The adulteration phenomenon is serious. Therefore, it is crucial to establish a fast and simple method in discriminating the natural, artificial and in vitro cultured Calculus bovis. Bile acids, one of the main active constituents, are taken as an important indicator for evaluating the quality of Calculus bovis and the substitutes. Several techniques have been built to analyze bile acids in Calculus bovis. Whereas, as bile acids are with poor ultraviolet absorbance and high structural similarity, effective technology for identification and quality control is still lacking. Methods: In this study, high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (LC/MS/MS) was applied in the analysis of bile acids, which effectively identified natural, artificial and in vitro cultured Calculus bovis and provide a new method for their quality control. Results: Natural, artificial and in vitro cultured Calculus bovis were differentiated by bile acids analysis. A new compound with protonated molecule at m/z 405 was found, which we called 3α, 12α-dihydroxy-7-oxo-5α-cholanic acid. This compound was discovered in in vitro cultured Calculus bovis, but almost not detected in natural and artificial Calculus bovis. A total of 13 constituents was identified. Among them, three bio-markers, including glycocholic acid, glycodeoxycholic acid and taurocholic acid (TCA) were detected in both natural and artificial Calculus bovis, but the density of TCA was different in two kinds of Calculus bovis. In addition, the characteristics of bile acids were illustrated. Conclusions: The HPLC coupled with tandem MS (LC/MS/MS) method was feasible, easy, rapid and accurate in
... it reduces the risk of food poisoning . Food irradiation is used in many countries. It was first approved in the U.S. to prevent sprouts on white potatoes, and to control insects on wheat and in certain spices and seasonings.
Martín-Atance, P; León-Vizcaíno, L; Palomares, F; Revilla, E; González-Candela, M; Calzada, J; Cubero-Pablo, M J; Delibes, M
We conducted a retrospective serologic survey for antibodies against the MPB70 protein of Mycobacterium bovis in wild carnivores from Doñana National Park (southwestern Spain). Serum samples from 118 red foxes (Vulpes vulpes), 39 Iberian lynx (Lynx pardinus), 31 Eurasian badgers (Meles meles), five Egyptian mongoose (Herpestes ichneumon), four European genet (Genetta genetta), and one Eurasian otter (Lutra lutra) were analyzed using an indirect competitive enzyme-linked immunoassay. Antibodies against the MPB70 protein of M. bovis were detected in seven badgers, five foxes, and one lynx. The frequency of positive animals was significantly higher in badger (23%) than in lynx (3%) and fox (4%). Antibodies were not detected in other species. Annual antibody frequency peaked at 38% in badgers and 11% for red fox. These species may contribute to persistence of bovine tuberculosis in Doñana.
Maragou, Chrysoula; Theologie-Lygidakis, Nadia; Ioannidis, Panayotis; Stenou, Antonia; Kanavaki, Sophia; Iatrou, Ioannis; Tsolia, Maria N
Bovine tuberculosis is a zoonotic disease, and although its incidence has dramatically decreased in developed countries where effective control measures are applied, it still remains a potential health hazard in the developing world. Tuberculosis of the oral cavity is extremely rare and is usually secondary to pulmonary involvement. We present the unusual case of an immunocompetent 6-year-old child residing in an urban area with primary oral tuberculosis due to Mycobacterium bovis, which was confirmed by the application of a molecular genetic approach. M. bovis belongs to Mycobacterium tuberculosis complex which comprises species with close genetic relationship, and for this reason, the use of new molecular techniques is a useful tool for the differentiation at species level of the closely related members of this complex.
Dekker, John P.
The Streptococcus bovis group has undergone significant taxonomic changes over the past 2 decades with the advent of new identification methods with higher discriminatory power. Although the current classification system is not yet embraced by all researchers in the field and debate remains over the performance of molecular techniques for identification to the species level within the group, important disease associations for several members of the group have been clarified. Here, we provide a brief overview of the history of the S. bovis group, an outline of the currently accepted classification scheme, a review of associated clinical syndromes, and a summary of the performance and diagnostic accuracy of currently available identification methods. PMID:26912760
Tomlinson, A J; Chambers, M A; Carter, S P; Wilson, G J; Smith, G C; McDonald, R A; Delahay, R J
The behaviour of certain infected individuals within socially structured populations can have a disproportionately large effect on the spatio-temporal distribution of infection. Endemic infection with Mycobacterium bovis in European badgers (Meles meles) in Great Britain and Ireland is an important source of bovine tuberculosis in cattle. Here we quantify the risk of infection in badger cubs in a high-density wild badger population, in relation to the infection status of resident adults. Over a 24-year period, we observed variation in the risk of cub infection, with those born into groups with resident infectious breeding females being over four times as likely to be detected excreting M. bovis than cubs from groups where there was no evidence of infection in adults. We discuss how our findings relate to the persistence of infection at both social group and population level, and the potential implications for disease control strategies.
De Castro, Cristina; Grice, I Darren; Daal, Terese-Marie; Peak, Ian R; Molinaro, Antonio; Wilson, Jennifer C
Moraxella bovis is a Gram-negative microorganism that causes Infectious Bovine Keratoconjunctivitis (IBK), colloquially known as 'Pink eye' in cattle worldwide. Lipopolysaccharides/lipooligosaccharides are the predominant glycans on the surface of Gram-negative microorganisms. Structural elucidation of the oligosaccharide structure of the rough phenotype of Moraxella bovis strain Epp63 was determined using GC-MS, methylation analysis, and NMR spectroscopy. The oligosaccharide is a branched structure that comprises 10 sugars in addition to KDO. The unusual features of this oligosaccharide include the fact that the oligosaccharide is devoid of heptose. The KDO residue is directly attached to a (→4,6)-branched glucose and additionally contains a terminal open chain acetal-linked N-acetylgalactosamine, (1S)-GalaNAc residue →4,6-linked to a sub-terminal galactose residue.
do Rosário, Tatiana Reis; Dib, Cristina Corsi; Roxo, Eliana; Pinheiro, Sônia Regina; Vasconcellos, Silvio Arruda; Benites, Nilson Roberti
The initial growth of mycobacteria from 49 samples of cattle and buffalo organs collected in commercial slaughterhouses was compared between modified Middlebrook 7H11 thin layer microcolony culture and Stonebrink medium used in the isolation of Mycobacterium bovis. Aliquots were decontaminated by Petroff’s method, processed and cultured in both media. The identity of the acid-fast bacilli stained by Ziehl-Neelsen was confirmed by PCR. Optical microscopy showed that results of the early observation of Mycobacterium bovis colonies in thin layer culture were similar to those obtained in macroscopic observation of the colonies in Stonebrink medium. However, early observation of the colonies enabled early confirmation by PCR, given the shorter time to the visualization of colonies when thin layer culture was used (between the 12nd and 25th day of culture). PMID:24948936
do Rosário, Tatiana Reis; Dib, Cristina Corsi; Roxo, Eliana; Pinheiro, Sônia Regina; Vasconcellos, Silvio Arruda; Benites, Nilson Roberti
The initial growth of mycobacteria from 49 samples of cattle and buffalo organs collected in commercial slaughterhouses was compared between modified Middlebrook 7H11 thin layer microcolony culture and Stonebrink medium used in the isolation of Mycobacterium bovis. Aliquots were decontaminated by Petroff's method, processed and cultured in both media. The identity of the acid-fast bacilli stained by Ziehl-Neelsen was confirmed by PCR. Optical microscopy showed that results of the early observation of Mycobacterium bovis colonies in thin layer culture were similar to those obtained in macroscopic observation of the colonies in Stonebrink medium. However, early observation of the colonies enabled early confirmation by PCR, given the shorter time to the visualization of colonies when thin layer culture was used (between the 12(nd) and 25(th) day of culture).
Lopez, A; Maxie, M G; Savan, M; Ruhnke, H L; Thomson, R G; Barnum, D A; Geissinger, H D
Based on current literature which commonly associates bovine virus diarrhea virus and Mycoplasma bovis with "pneumonic pasteurellosis," an investigation was conducted into the effect of these two pathogens on the capacity of bovine lung to clear inhaled Pasteurella haemolytica. There was no significant effect (p less than 0.05) of either bovine virus diarrhea virus or M. bovis on the mean clearance rate of P. haemolytica, nor did the time interval of three, five or seven days between the first inoculation and exposure to P. haemolytica and adversely affect the lung clearance rates. However, it was found that the left lungs and a higher bacterial retention (p less than 0.05) than the right lungs. PMID:7127194
Zaugg, J L; Lane, V M
We evaluated the efficacy of buparvaquone in eliminating infection with Babesia equi of European origin in carrier horses and in splenectomized horses with experimentally induced acute infection. When administered at the rate of 5 mg/kg of body weight, IV, 4 times at 48-hour intervals, buparvaquone prompted rapid abatement of parasitemia. However, secondary and tertiary recrudescent parasitemias invariably returned with establishment of the carrier state. Buparvaquone, at the dosage evaluated, had transitory therapeutic efficacy against acute B equi infection in splenectomized horses, but was unable alone to clear carrier infection.
Taubert, Anja; Wimmers, Klaus; Ponsuksili, Siriluck; Jimenez, Cristina Arce; Zahner, Horst; Hermosilla, Carlos
Within its life cycle Eimeria bovis undergoes a long lasting intracellular development into large macromeronts in endothelial cells. Since little is known about the molecular basis of E. bovis-triggered host cell regulation we applied a microarray-based approach to define transcript variation in bovine endothelial cells early after sporozoite invasion (4 h post inoculation (p.i.)), during trophozoite establishment (4 days p.i.), during early parasite proliferation (8 days p.i.) and towards macromeront maturation (14 days p.i.). E. bovis infection led to significant changes in the abundance of many host cell gene transcripts. As infection progressed, the number of regulated genes increased such that 12, 45, 175 and 1184 sequences were modulated at 4 h, 4, 8 and 14 days p.i., respectively. These genes significantly interfered with several host cell functions, networks and canonical pathways, especially those involved in cellular development, cell cycle, cell death, immune response and metabolism. The correlation between stage of infection and the number of regulated genes involved in different aspects of metabolism suggest parasite-derived exploitation of host cell nutrients. The modulation of genes involved in cell cycle arrest and host cell apoptosis corresponds to morphological in vitro findings and underline the importance of these aspects for parasite survival. Nevertheless, the increasing numbers of modulated transcripts associated with immune responses also demonstrate the defensive capacity of the endothelial host cell. Overall, this work reveals a panel of novel candidate genes involved in E. bovis-triggered host cell modulation, providing a valuable tool for future work on this topic.
Bigi, F; Alito, A; Fisanotti, J C; Romano, M I; Cataldi, A
Serum from naturally infected cattle was used to identify a novel Mycobacterium bovis antigen from an expression library. The first recombinant product identified was a fusion protein with lacZ (55 kDa). A clone containing the whole gene was also obtained. This clone expressed a 38-kDa protein. A rabbit serum against the recombinant antigen reacts in M. bovis supernatants with two proteins of 36 and 34 kDa. The new protein was called P36/P34. The gene cloned has a deduced amino acid sequence with a predicted molecular mass of 28 kDa, showing a characteristic signal sequence for exportation. The protein bears partial homology to a 28-kDa protein from M. leprae. An interesting feature of the P36/P34 sequence is that it contains several PGLTS repeats, which are not present in the M. leprae protein. Antigenic determinants seem also to be conserved between the two proteins because sera from leprosy patients recognized the recombinant M. bovis protein. The discrepancy among the molecular mass deduced from the sequence (28 kDa), that of the recombinant protein in Escherichia coli (38 kDa), and that of the native protein in M. bovis (36 and 34 kDa) could be attributed to posttranslational modifications or to the high proline content that may alter the migration properties of the protein. This antigen seems to be immunodominant during bovine tuberculosis, because 8 of 9 serum specimens from diseased cattle are reactive. The homology among the M. leprae 28-kDa protein, the protein described in this article, and a recently described M. tuberculosis protein suggests the existence of a new protein family in mycobacteria.
Farrell, Damien; Stuber, Tod P.; Schubert, Olga T.; Aebersold, Ruedi; Robbe-Austerman, Suelee
ABSTRACT We report here an update to the reference genome sequence of the bovine tuberculosis bacillus Mycobacterium bovis AF2122/97, generated using an integrative multiomics approach. The update includes 42 new coding sequences (CDSs), 14 modified annotations, 26 single-nucleotide polymorphism (SNP) corrections, and disclosure that the RD900 locus, previously described as absent from the genome, is in fact present. PMID:28385856
Nava Vargas, Alejandro; Milián Suazo, Feliciano; Cantó Alarcón, Germinal Jorge; Rubio Venegas, Yezenia; Guerrero Solorio, Roberto; Rodríguez Hernández, Elba; Pizano Martìnez, Oscar
Bovine tuberculosis (bTB) is a disease caused by Mycobacterium bovis (M. bovis), which affects cattle, animal species and humans. To determinate the genetic structure of strains of M. bovis in mexican cattle, 467 isolates obtained from 2009 to 2010 from different regions of Mexico with known spoligotype were included in the study. The isolates were genotyped by interspersed repeated mycobacterial units-variable number tandem repeats (MIRU-VNTR) obtaining 13 MIRU-VNTR groups. When combining MIRU-VNTR patterns with its spolygotypes, the Hunter genetic discrimination index (HGDI), we obtained 421 genetic patterns distributed in 17 groups. The HGDI for the total loci was 0.99. The locus that presented the higher HGDI was 2461 (0.857), while the locus with the lowest HGDI was 2686 (0.239). When we analyzed our results, using just 6 or 8 MIRU-VNTR we obtained an discriminatory power of 0.8499 and 0.8875 respectively indicating lower HGDI than 12 MIRU-VNTR locus.
Lesellier, Sandrine; Corner, Leigh; Costello, Eamon; Sleeman, Paddy; Lyashchenko, Konstantin; Greenwald, Rena; Esfandiari, Javan; Singh, Mahavir; Hewinson, R Glyn; Chambers, Mark; Gormley, Eamonn
European badgers (Meles meles) are considered to be an important reservoir of infection for Mycobacterium bovis and are implicated in the transmission of tuberculosis to cattle in Ireland and Great Britain. Accurate tests are required for tuberculosis surveillance in badger populations and to provide a basis for the development of strategies, including vaccination, to reduce the incidence of the infection. In this study, we have developed an endobronchial M. bovis infection model in badgers in which we measured cell-mediated immune and serological responses for up to 24 weeks post-infection. Groups of badgers were subjected to necropsy at 6-week intervals and the gross lesion severity status compared with immune responses measured in blood samples taken throughout the course of the study. The panel of antigens included bovine and avian tuberculins (PPD) as well as single antigens, ESAT-6, CFP-10, MPB70, Rv3019c, Rv3873, Rv3878 and Rv3879, all known to be recognised by the immune system in other animal models of tuberculosis infection. Our results demonstrated that M. bovis infected badgers responded to specific antigens as early as 6 weeks post-infection, consistent with the presence of visible lesions. The data also revealed unique patterns of antigen recognition with high levels of PBMC proliferation in the presence of CFP-10 but low proliferation levels with ESAT-6. Using a multi-antigen print immunoassay (MAPIA), we were able to confirm that MPB83 is the dominant antigen recognised by serum antibodies in infected badgers.
Lesellier, Sandrine; Corner, Leigh; Costello, Eamon; Lyashchenko, Konstantin; Greenwald, Rena; Esfandiari, Javan; Singh, Mahavir; Hewinson, R Glyn; Chambers, Mark; Gormley, Eamonn
European badgers (Meles meles) are a reservoir host of Mycobacterium bovis and are implicated in the transmission of tuberculosis to cattle in Ireland and Great Britain. The development of a vaccine for use in badgers is considered a key element of any campaign to eradicate the disease in livestock in both countries. In this study we have vaccinated groups of badgers with approximately 5 x 10(5)cfu of the BCG vaccine delivered via two alternative routes, subcutaneous and mucosal (intranasal/conjunctival). Following experimental endobronchial infection with approximately 10(4)cfu of M. bovis, all badgers were euthanised at 12 weeks post-infection. At post-mortem examination both vaccinated groups had significantly reduced severity of disease compared with the non-vaccinated controls. The analysis of immune responses throughout the study showed that vaccination with BCG did not generate any detectable immunological responses as measured by IFN-gamma production in antigen-stimulated peripheral blood mononuclear cells (PBMC) and IgG serological responses. However, the levels of the responses increased following M. bovis infection, and the kinetic profiles corresponded to the severity of lesions recorded post-mortem. Significant differences were observed in the timing of development of the immune responses between vaccinates and controls. The results suggest that the immunological responses are associated with the levels of protective immunity and could be used as markers to monitor control of disease in badgers following vaccination.
Palmer, Mitchell V.; Thacker, Tyler C.; Waters, W. Ray; Gortázar, Christian; Corner, Leigh A. L.
Complex and dynamic interactions involving domestic animals, wildlife, and humans create environments favorable to the emergence of new diseases, or reemergence of diseases in new host species. Today, reservoirs of Mycobacterium bovis, the causative agent of tuberculosis in animals, and sometimes humans, exist in a range of countries and wild animal populations. Free-ranging populations of white-tailed deer in the US, brushtail possum in New Zealand, badger in the Republic of Ireland and the United Kingdom, and wild boar in Spain exemplify established reservoirs of M. bovis. Establishment of these reservoirs is the result of factors such as spillover from livestock, translocation of wildlife, supplemental feeding of wildlife, and wildlife population densities beyond normal habitat carrying capacities. As many countries attempt to eradicate M. bovis from livestock, efforts are impeded by spillback from wildlife reservoirs. It will not be possible to eradicate this important zoonosis from livestock unless transmission between wildlife and domestic animals is halted. Such an endeavor will require a collaborative effort between agricultural, wildlife, environmental, and political interests. PMID:22737588
Pugh, G W; Wilbur, L A; Beall, C W
Selected cultures of Moraxella bovis were studied in calves, using chambers fabricated from a semipermeable membrane supported and protected by perforated plastic golf balls. Plain balls, semipermeable membrane-covered balls, and balls that contained bags fabricated from semipermeable membranes were surgically implanted subcutaneously in calves at a site approximately 25 cm ventral to the paralumbar fossa and cranial to the prefemoral lymph node. After calves recovered from surgery, chambers were inoculated with different cultures of M bovis. Cultural examination of samples taken from inoculated chambers indicated that M bovis cultures were maintained within the chambers for variable times, depending on the characteristics of the culture inoculated. A smooth (piliated) culture dissociated into a rough (nonpiliated) culture after 4 weeks of incubation within a chamber but none of the rough cultures became smooth. The technique offers a method to study the in vivo variations in a series of cultures or strains of microorganisms under the influence of the same host factors. The selective permeability of the chamber may be controlled by using membranes of different porosities to control the flow of entering or exiting dialysate.
King, Hayley C; Murphy, Andrew; James, Phillip; Travis, Emma; Porter, David; Hung, Yu-Jiun; Sawyer, Jason; Cork, Jennifer; Delahay, Richard J; Gaze, William; Courtenay, Orin; Wellington, Elizabeth M
The incidence of Mycobacterium bovis, the causative agent of bovine tuberculosis, has been increasing in UK cattle herds resulting in substantial economic losses. The European badger (Meles meles) is implicated as a wildlife reservoir of infection. One likely route of transmission to cattle is through exposure to infected badger urine and faeces. The relative importance of the environment in transmission remains unknown, in part due to the lack of information on the distribution and magnitude of environmental reservoirs. Here we identify potential infection hotspots in the badger population and quantify the heterogeneity in bacterial load; with infected badgers shedding between 1 × 10(3)- 4 × 10(5) M. bovis cells g(-1) of faeces, creating a substantial and seasonally variable environmental reservoir. Our findings highlight the potential importance of monitoring environmental reservoirs of M. bovis which may constitute a component of disease spread that is currently overlooked and yet may be responsible for a proportion of transmission amongst badgers and onwards to cattle.
Sharma, Shukriti; Markham, Philip F.; Browning, Glenn F.
Mycoplasmas are regarded to be useful models for studying the minimum genetic complement required for independent survival of an organism. Mycoplasma bovis is a globally distributed pathogen causing pneumonia, mastitis, arthritis, otitis media and reproductive tract disease, and genome sequences of three strains, the type strain PG45 and two strains isolated in China, have been published. In this study, several Tn4001 based transposon constructs were generated and used to create a M. bovis PG45 insertional mutant library. Direct genome sequencing of 319 independent insertions detected disruptions in 129 genes in M. bovis, 48 of which had homologues in Mycoplasma mycoides subspecies mycoides SC and 99 of which had homologues in Mycoplasma agalactiae. Sixteen genes found to be essential in previous studies on other mycoplasma species were found to be dispensable. Five of these genes have previously been predicted to be part of the core set of 153 essential genes in mycoplasmas. Thus this study has extended the list of non-essential genes of mycoplasmas from that previously generated by studies in other species. PMID:24897538
Shimizu, Ernesto; Macías, Analía; Paolicchi, Fernando; Magnano, Gabriel; Zapata, Laura; Fernández, Analía; Canal, Ana; Garbaccio, Sergio; Cataldi, Angel; Caimi, Karina; Zumárraga, Martín
Mycobacterium bovis is the causative agent of bovine tuberculosis (TB), a disease that affects approximately 5% of Argentinean cattle. Among the molecular methods for genotyping, the most convenient are spoligotyping and variable number of tandem repeats (VNTR). A total of 378 samples from bovines with visible lesions consistent with TB were collected at slaughterhouses in three provinces, yielding 265 M. bovis spoligotyped isolates, which were distributed into 35 spoligotypes. In addition, 197 isolates were also typed by the VNTR method and 54 combined VNTR types were detected. There were 24 clusters and 27 orphan types. When both typing methods were combined, 98 spoligotypes and VNTR types were observed with 27 clusters and 71 orphan types. By performing a meta-analysis with previous spoligotyping results, we identified regional and temporal trends in the population structure of M. bovis. For SB0140, the most predominant spoligotype in Argentina, the prevalence percentage remained high during different periods, varying from 25.5-57.8% (1994-2011). By contrast, the second and third most prevalent spoligotypes exhibited important fluctuations. This study shows that there has been an expansion in ancestral lineages as demonstrated by spoligotyping. However, exact tandem repeat typing suggests dynamic changes in the clonal population of this microorganism. PMID:24676658
Sühwold, Anke; Hermosilla, Carlos; Seeger, Torsten; Zahner, Horst; Taubert, Anja
Eimeria bovis infections commonly have clinical impact only on young animals, as homologous reinfections generally are under immunological control. So far, the nature of the immune responses delivering protection to calves has not been investigated. In this study we therefore analysed local and peripheral proliferative T cell activities of primary and challenge-infected calves and investigated the occurrence of T cell phenotypes in the peripheral blood and in mucosal gut segments isolated either by bioptic means or by necropsies.We show that lymphocytes of E. bovis-infected calves exhibit effective, transient antigen-specific proliferative responses in the course of prepatency of primary infection but fail to react after homologous reinfection suggesting early abrogation of parasite development. Whilst in primary infection an expansion of peripheral CD4+ T cells was observed, reinfection had no effect on the proportions of CD4+, CD8+ subsets or gammadeltaTCR+ T cells. In contrast, both E. bovis primary and challenge infections had an impact on local tissue T cell distribution. Primary infection was characterised by a CD4+ T cell infiltration early in prepatency in ileum and later in colon mucosa, whereas CD8+ T cells were only found accumulating in the latter gut segment. Challenge infection led to infiltration of both CD4+ and CD8+ T cells in small intestine and large intestine segments indicating protective functions of both cell types. In contrast, infiltration of ileum and colon mucosa with gammadeltaTCR+ T cells was restricted to primary infection.
de Lisle, Geoffrey W; Kawakami, R Pamela; Yates, Gary F; Collins, Desmond M
As part of wildlife surveillance for bovine tuberculosis, pooled lymph nodes from 21,481 ferrets, 1056 stoats and 83 weasels were cultured for mycobacteria. A total of 268 isolates of Mycobacterium bovis were obtained from ferrets, 2 from stoats and none from weasels, demonstrating the presence of a wildlife reservoir of infection in ferrets. DNA typing by restriction endonuclease analysis (REA) of 48 selected isolates of M. bovis revealed 23 REA types. Twenty-one of these types had previously been isolated from cattle and farmed deer, demonstrating a complex cycle of infection involving wildlife and domestic animals. Apart from M. bovis, a further 208 mycobacterial isolates were obtained, the majority of which (178) were members of the M. avium complex. Speciation of the remaining 30 mycobacterial isolates by DNA sequencing of the 16s rRNA gene, identified half the isolates as M. triplex. Other species identified included M. fortuitum, M. florentinum, M. interjectum, M. intracellulare, M. holsaticum, and M. septicum/M. peregrinum.
Zumárraga, Martín José; Soutullo, Adriana; García, María Inés; Marini, Rocío; Abdala, Alejandro; Tarabla, Héctor; Echaide, Susana; López, Marcela; Zervini, Elsa; Canal, Ana; Cataldi, Angel Adrián
Bovine tuberculosis (bTB) is a chronic and zoonotic disease due to Mycobacterium bovis. The tuberculosis eradication campaign carried out in Argentina has considerably improved the health situation of the herds. Here we evaluated a strategy to detect M. bovis-infected herds by Touch-Down IS6110 polymerase chain reaction (PCR) in bulk tank raw milk from dairy farms. We evaluated 177 samples from herds with the official tuberculosis free certificate (TFC) and 80 from herds without the certificate, non-tuberculosis-free certificate (NTFC), from 10 departments of Santa Fe province, Argentina. To avoid the effect of Taq polymerase inhibitors, a dilution of DNA template was performed. Positive PCR results were obtained in 102 (40%) of the samples, whereas negative ones were obtained in 155 (60%) of the samples. Importantly, 44% of NTFC and 38% of TFC samples were positive. All samples were subjected to culture in Löwenstein Jensen and Stonebrink media with no positive isolation. The negative predictive value (NPV) of PCR in the TFC group was 95%, while the positive predictive value (PPV) of PCR in the NTFC group was 51%. Based on these results, this work proposes a method that should be applied regularly to detect M. bovis--infected dairy herds, complementary to the official test of tuberculin, or purifed protein derivative (PPD), to control dairy herds, especially those free of tuberculosis.
Gerchman, Irena; Levisohn, Sharon; Mikula, Inna; Lysnyansky, Inna
Monitoring of susceptibility to antibiotics in field isolates of pathogenic bovine mycoplasmas is important for appropriate choice of treatment. Our study compared in vitro susceptibility profiles of Mycoplasma bovis clinical strains, isolated during 2005-2007 from Israeli and imported calves. Minimal inhibitory concentration (MIC) values were determined for macrolides by the microbroth dilution test, for aminoglycosides by commercial Etest, and for fluoroquinolones and tetracyclines by both methods. Notably, although correlation between the methods was generally good, it was not possible to determine the MIC endpoint for enrofloxacin-resistant strains (MIC > or =2.5 microg/ml in the microtest) by Etest. Comparison of antibiotic susceptibility profiles between local and imported M. bovis strains revealed that local strains were significantly more resistant to macrolides than most isolates from imported animals, with MIC(50) of 128 microg/ml vs. 2 microg/ml for tilmicosin and 8 microg/ml vs. 1 microg/ml for tylosin, respectively. However, local strains were more susceptible than most imported strains to fluoroquinolones and spectinomycin. Difference in susceptibility to tetracycline, doxycycline and oxytetracycline between local and imported strains was expressed in MIC(90) values for imported strains in the susceptible range compared to intermediate susceptibility for local strains. The marked difference in susceptibility profiles of M. bovis strains isolated from different geographical regions seen in this study emphasizes the necessity for performing of the antimicrobial susceptibility testing periodically and on a regional basis.
Shimizu, Ernesto; Macías, Analía; Paolicchi, Fernando; Magnano, Gabriel; Zapata, Laura; Fernández, Analía; Canal, Ana; Garbaccio, Sergio; Cataldi, Angel; Caimi, Karina; Zumárraga, Martín
Mycobacterium bovis is the causative agent of bovine tuberculosis (TB), a disease that affects approximately 5% of Argentinean cattle. Among the molecular methods for genotyping, the most convenient are spoligotyping and variable number of tandem repeats (VNTR). A total of 378 samples from bovines with visible lesions consistent with TB were collected at slaughterhouses in three provinces, yielding 265 M. bovis spoligotyped isolates, which were distributed into 35 spoligotypes. In addition, 197 isolates were also typed by the VNTR method and 54 combined VNTR types were detected. There were 24 clusters and 27 orphan types. When both typing methods were combined, 98 spoligotypes and VNTR types were observed with 27 clusters and 71 orphan types. By performing a meta-analysis with previous spoligotyping results, we identified regional and temporal trends in the population structure of M. bovis. For SB0140, the most predominant spoligotype in Argentina, the prevalence percentage remained high during different periods, varying from 25.5-57.8% (1994-2011). By contrast, the second and third most prevalent spoligotypes exhibited important fluctuations. This study shows that there has been an expansion in ancestral lineages as demonstrated by spoligotyping. However, exact tandem repeat typing suggests dynamic changes in the clonal population of this microorganism.
Fitzgerald, Scott D; Sledge, Dodd G; Maes, Roger; Wise, Annabel; Kiupel, Matti
Bovine leukosis associated with infection with the delta retrovirus Bovine leukemia virus (BLV) is endemic in many cattle herds in the United States. Infection has been associated with immunosuppression and decreased productivity. Cases of tuberculosis in cows due to infection with Mycobacterium bovis reemerged in Michigan in 1998, and despite intensive eradication attempts, new cases of bovine tuberculosis are sporadically identified. The present report details a coinfection with BLV and M. bovis in a Holstein cow from Michigan that presented as part of a bovine tuberculosis screening program. Peripheral and visceral lymph nodes of this animal were markedly enlarged, homogeneously pale white, and bulged on the cut surface. The submandibular, mesenteric, and caudal mediastinal lymph nodes contained multifocal to coalescing caseogranulomas that ranged from 1 to 5 cm in diameter. Histologically, dense sheets of monomorphic populations of neoplastic lymphocytes obliterated the normal architecture of all lymph nodes. Caseogranulomas were characterized by central pools of amorphous degenerate eosinophilic and occasionally mineralized granular debris surrounded by thick rims of epithelioid macrophages, occasional Langhan's type giant cells, and fibrosis. Polymerase chain reaction assay was positive for BLV. Cultures of affected lymph nodes yielded growth of M. bovis.
Corner, L A L; Costello, E; Lesellier, S; O'Meara, D; Sleeman, D P; Gormley, E
The aim was to develop an endobronchial infection procedure for the study of Mycobacterium bovis infection in badgers. The badgers were anaesthetised and a cannula was passed per os to the tracheal bifurcation. When in place 1 ml of M. bovis suspension was inoculated. Three concentrations of M. bovis suspension were used; <10 colony forming units (cfu), approximately 10(2) cfu and approximately 3 x 10(3) cfu. The badgers were examined at three weekly intervals for clinical signs of disease and a tracheal aspirate was collected at each examination. The badgers were euthanased 17 weeks post infection (pi) and at the post mortem examination a wide range of tissues were examined for gross and histopathological lesions of tuberculosis and cultured for M. bovis. A sample of bronchial alveolar lavage (BAL) fluid was collected at post mortem for culture. At post mortem examination 17 weeks after infection, gross and histopathological lesions of tuberculosis were observed in all badgers inoculated with the high and medium dose and 1/3 inoculated with the low dose. M. bovis was recovered from all inoculated badgers. Infection in the high dose group was more widely disseminated than in the other groups. The number of sites with gross and histopathological lesions increased with increasing dose of M. bovis. All tracheal aspirates were negative on culture and only one BAL, collected from a badger of the high dose group, was positive on culture. No clinical signs due to the experimental infection were observed. The endobronchial route of inoculation is an effective route for establishing experimental infection, and could be used for studies of tuberculosis pathogenesis, immunology of M. bovis infection in badgers and for challenging badgers in vaccine protection studies. Badgers appeared to be very susceptible to infection by this procedure even with a dose of < 10 cfu but appear to control and limit the resulting infection.
Martínez-Neri, Priscila A; López-Rincón, Gonzalo; Mancilla-Jiménez, Raúl; del Toro-Arreola, Susana; Muñoz-Valle, José Francisco; Fafutis-Morris, Mary; Bueno-Topete, Miriam Ruth; Estrada-Chávez, Ciro; Pereira-Suárez, Ana Laura
The immunomodulatory functions of prolactin (PRL) are well recognized. Augmented PRL plasma levels were observed in patients with advanced tuberculosis (TB). Recently, we have reported that LPS and Mycobacterium bovis (M. bovis) induced differential expression of PRL receptor (PRLR) isoforms in THP-1 cells and bovine macrophages, respectively. The aim of this work was to determine whether PRL should be considered as a potential modulator of the signaling pathways and cytokine synthesis, induced by culture filtrate protein (CFP) from M. bovis in THP-1 monocytes. The THP-1 cells were stimulated with PRL (20ng/mL), M. bovis CFP (50μg/mL). PRLR as well as phosphorylated STAT3, STAT5, Akt1/2/3, ERK1/2 and p38 expression were evaluated by Western blot. IL1-β, TNF-α, IL-6, IL-12, IL-8, and IL-10 concentrations were measured by ELISA. Our results demonstrated that the expression pattern of PRLR short isoforms is induced by M. bovis CFP. M bovis CFP induced phosphorylation of Akt2, ERK1/2, p38, STAT3, and STAT5 pathways. In turn, PRL only activated the JAK2/STAT3-5 signaling pathway. However, when combined both stimuli, PRL significantly increased STAT3-5 phosphorylation and downregulated Akt2, ERK1/2, and p38 phosphorylation. As expected, M. bovis CFP induced substantial amounts of IL1-β, IL-6, TNF-α, IL-8, IL-12, and IL-10. However, the PRL costimulation considerably decreased IL1-β, TNF-α, and IL-12 secretion, and increased IL-10 production. This results suggest that up-regulation of IL-10 by PRL might be modulating the pro-inflammatory response against mycobacterial antigens through the MAPK pathway.
Rapoport, Adi; Aharonson-Raz, Karin; Berlin, Dalia; Tal, Saar; Gottlieb, Yuval; Klement, Eyal; Steinman, Amir
Equine piroplasmosis imposes great concerns for the equine industry regarding international horse movement, and therefore requires reliable diagnostic tools. Recent studies from South Africa and Jordan, including a preliminary study in Israel, reported extremely low seroprevalence to Babesia caballi (B. caballi) (0-1%) using the acceptable rhoptry-associated protein-1 (RAP-1) cELISA. In accordance with the study from South Africa demonstrating a significant heterogeneity in the rap-1 gene sequence of South African B. caballi isolates, the objectives of this study were to phylogenetically characterize the rap-1 gene of the Israeli isolates and determine the prevalence of B. caballi in horses in Israel. Out of 273 horses tested using the RAP-1 cELISA, only one was sero-positive, while 9.3% were positive on PCR performed on the rap-1 gene. Phylogenetic analysis of the rap-1 gene grouped the Israeli isolates in a cluster together with the South African strains (99% nt identity), but in a separate cluster from the American/Caribbean strains (81-82% nt identity). These findings support the existence of heterogeneity in the RAP-1 amino-acid sequences of the Israeli and South African isolates as compared to that used in the cELISA commercial kit and raise doubts as to the ability of this assay to serve as a sole regulatory test for international horse movement. Risk factor analysis found management and age to significantly associate with prevalence of B. caballi, as higher prevalence was noted in horses held out on pasture and a negative association was recorded with age. In addition, B. caballi was not detected in horses in the steppe-arid and extreme-arid climatic regions as compared to the wetter regions. Findings of this study emphasize the need to combine several detection methods to ameliorate the control and spread of the disease.
Yamasaki, Masahiro; Harada, Eriko; Tamura, Yu; Lim, Sue Yee; Ohsuga, Tatsuyuki; Yokoyama, Nozomu; Morishita, Keitaro; Nakamura, Kensuke; Ohta, Hiroshi; Takiguchi, Mitsuyoshi
Babesia gibsoni is a causative pathogen of canine babesiosis, which is commonly treated with anti-babesial drugs; however, the development of novel, more effective anti-babesial drugs is necessary because the currently used drugs cannot remove the parasites from dogs. Therefore we investigated the anti-babesial effect of amphotericin B (AmB), a membrane-active polyene macrolide antibiotic. The interaction of such compounds with sterols in bilayer cell membranes can lead to cell damage and ultimately cell lysis. AmB exhibits in vitro activity against B. gibsoni in normal canine erythrocytes within 12h. We also studied liposomal AmB (L-AmB), a liposomal formulation of AmB that required a longer incubation period to reduce the number of parasites. However, L-AmB completely inhibited the invasion of free parasites into erythrocytes. These results indicated that free parasites failed to invade erythrocytes in the presence of L-AmB. Both AmB and L-AmB induced mild hemolysis of erythrocytes. Moreover, the methemoglobin level and the turbidity index of erythrocytes were significantly increased when erythrocytes were incubated with AmB, suggesting that AmB induced oxidative damage in erythrocytes. Finally, the anti-babesial activity of AmB in vivo was observed. When experimentally B. gibsoni-infected dogs were administered 0.5 and 1mg/kg AmB by the intravenous route, the number of parasites decreased; however, recurrence of parasitemia was observed, indicating that AmB did not eliminate parasites completely. Blood urea nitrogen and creatinine of dogs were abnormally elevated after the administration of 1mg/kg AmB. These results indicate that AmB has in vivo activity against B. gibsoni; however, it does not eliminate parasites from infected dogs and affects kidney function at a high dose.
Camacho, Angel Tomas; Guitian, Francisco Javier; Pallas, Estrella; Gestal, Juan Jesus; Olmeda, Sonia; Goethert, Heidi; Telford, Sam; Spielman, Andrew
Babesia annae piroplasms have recently been recognised as a cause of infection and disease among dogs in Europe. The pathogenesis and clinical implications of this emerging disease remain poorly understood. We conducted this study to describe the electrophoretic profiles associated with the infection and to determine if B. annae associated azotaemia is caused by renal failure. We examined by microscopy 2,979 canine blood samples submitted to a diagnostic laboratory in NW Spain between September 2001 and April 2002. Small ring-shaped piroplasms were detected in blood smears of 87 samples and the identity of 58 of these presumptive cases were confirmed by PCR. This group of 58 infected dogs and a reference group of 15 healthy non-infected dogs were our study population. For all the dogs, serum protein response to -albumin, alpha-1 globulin, alpha-2 globulin, beta globulin and gamma globulin- was measured by capillary electrophoresis. The response of infected and non-infected dogs was compared and within infected dogs, the response of those with azotaemia (19) was compared with that of non-azotaemic dogs (39). Infected dogs presented a significant elevation of total proteins and all the different globulin fractions, and significantly lower levels of albumin compared to non-infected dogs. Among infected dogs, those presenting azotaemia had significantly lower concentrations of total proteins, albumin, beta and gamma globulins, and significantly higher values of alpha-2 globulin. Specific gravity was below the threshold of 1,025 for all dogs with azotaemia for which a urine sample was available (7) suggesting that azotaemia, in these dogs was of renal origin. Azotaemic dogs had higher concentrations of cholesterol and triglycerides, probably as a result of a liver compensatory response to the loss of proteins. We conclude that serum protein response in B. annae infected dogs corresponds to the pattern of a haemolytic syndrome with intense inflammatory reaction and that
Wesonga, F D; Gachohi, J M; Kitala, P M; Gathuma, J M; Njenga, M J
Anaplasma marginale and Babesia bigemina are important tick-borne pathogens of cattle. A cross-sectional survey was undertaken to determine the seroprevalence of A. marginale and B. bigemina infections and identify associated risk factors on traditional smallholder farms in Machakos County, Kenya. A total of 421 cattle from 127 farms from four divisions in the county were sampled and visited between September and November 2007. The farms were selected by a proportional allocation approach based on the number of farms in the four divisions previously selected by stratified random sampling method. Information on animal and individual farm management variables was obtained using standardized questionnaires. Prevalence of serum antibodies due to A. marginale and B. bigemina pathogens was determined using the enzyme-linked immunosorbent assay (ELISA) technique. The relationship between the seropositivity and associated risk factors was assessed by multivariable analyses using standard logistic regression models. The overall estimation (and their 95% confidence intervals) of A. marginale and B. bigemina seropositivity at the animal level was 53.4% (48.5%, 58.2%) and 40.6% (35.8%, 45.4%), respectively. Two variables, "animal age" and "administrative division," were significantly associated with the A. marginale seroresponse. Three variables, "animal age" "grazing system" and "administrative division" were significantly associated with the B. bigemina seroresponse. These findings suggest possible indicators of existence of endemic instability for the two infections. The study identifies characterization of environmental suitability for the vectors and how they interact with grazing systems to cause the infections as an area for further studies, for improved understanding of the infections and in designing disease control programs.
Yamane, I; Thomford, J W; Gardner, I A; Dubey, J P; Levy, M; Conrad, P A
We determined the extent of the serologic cross-reactivity in the indirect fluorescent antibody (IFA) test for Babesia gibsoni, and the optimal cut-off titer for seropositivity in the test. The lowest titer to B gibsoni detected in a dog with naturally acquired clinical babesiosis was 1,280, and 7 of 12 dogs had titer between 10,240 and 20,480. Two experimentally infected normosplenic dogs developed high titer (40,960 to 81,920) to B gibsoni, and the same sera reacted in IFA tests for B canis (titer < or = 640), Toxoplasma gondii (titer < or = 2,560), and Neospora caninum (titer < or = 10,240). Dogs that were experimentally infected with B canis and T gondii had titer < or = 160 to B gibsoni. Dogs that were experimentally infected with N caninum had titer (80 to 10,240) to N caninum, but failed to have serologic reactivity to B gibsoni. Serologic titer of healthy dogs from Australia, a country where B gibsoni is not known to exist, was < or = 160 to B gibsoni. On the basis of these findings, a cut-off titer of 320 was considered to be appropriate for serodiagnosis of B gibsoni in dogs with clinical signs of babesiosis. A more conservative titer of 1,280 was established as the cut-off titer for seroepidemiologic studies based on relative costs and benefits of false-positive results and failure to isolate B gibsoni parasites after splenectomy and immunosuppression from a clinically normal dog with B gibsoni titer of 640.(ABSTRACT TRUNCATED AT 250 WORDS)
Terkawi, Mohamad Alaa; Cao, Shinuo; Herbas, Maria S; Nishimura, Maki; Li, Yan; Moumouni, Paul Franck Adjou; Pyarokhil, Asadullah Hamid; Kondoh, Daisuke; Kitamura, Nobuo; Nishikawa, Yoshifumi; Kato, Kentaro; Yokoyama, Naoaki; Zhou, Jinlin; Suzuki, Hiroshi; Igarashi, Ikuo; Xuan, Xuenan
In the present study, we examined the contributions of macrophages to the outcome of infection with Babesia microti, the etiological agent of human and rodent babesiosis, in BALB/c mice. Mice were treated with clodronate liposome at different times during the course of B. microti infection in order to deplete the macrophages. Notably, a depletion of host macrophages at the early and acute phases of infection caused a significant elevation of parasitemia associated with remarkable mortality in the mice. The depletion of macrophages at the resolving and latent phases of infection resulted in an immediate and temporal exacerbation of parasitemia coupled with mortality in mice. Reconstituting clodronate liposome-treated mice at the acute phase of infection with macrophages from naive mice resulted in a slight reduction in parasitemia with improved survival compared to that of mice that received the drug alone. These results indicate that macrophages play a crucial role in the control of and resistance to B. microti infection in mice. Moreover, analyses of host immune responses revealed that macrophage-depleted mice diminished their production of Th1 cell cytokines, including gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α). Furthermore, depletion of macrophages at different times exaggerated the pathogenesis of the infection in deficient IFN-γ(-/-) and severe combined immunodeficiency (SCID) mice. Collectively, our data provide important clues about the role of macrophages in the resistance and control of B. microti and imply that the severity of the infection in immunocompromised patients might be due to impairment of macrophage function.
Background Canine babesiosis due to Babesia canis is an endemic disease in many European countries. A vaccine is available in some countries, but it does not prevent the infection and just helps in reducing the gravity of clinical signs. Therefore, the major way to help preventing the disease is by controlling tick infestations on dogs. To assess the preventive efficacy of afoxolaner (NexGard®), a new oral anti- flea and tick product, against Babesia canis infected adult Dermacentor reticulatus in an experimentally controlled study. Methods Sixteen healthy mixed breed adult dogs, negative for Babesia canis antibodies were included in a single centre, randomized, blinded and controlled study to evaluate the impact of treatment with afoxolaner on the transmission of Babesia canis to dogs exposed to Dermacentor reticulatus. The dogs were randomly allocated into two groups of 8 dogs each. One group remained untreated. In the other group, dogs were treated orally with a novel formulation of afoxolaner (NexGard®) on day 0. All dogs were infested each by 50 adult Dermacentor reticulatus ticks (equal sex ratio) at days 7, 14, 21 and 28. The Dermacentor reticulatus ticks were confirmed to harbour Babesia canis by Polymerase Chain Reaction (PCR). Results The treatment was well tolerated by all dogs without any adverse effects. Babesia canis was transmitted by D. reticulatus to all untreated control dogs, confirmed following demonstration of hyperthermia, detection of B. canis parasites in blood smears and PCR assay from blood and serology. These confirmed infected dogs were subsequently treated with imidocarb and diminazene. The treated dogs remained negative based on all criteria until the last study, Day 56, confirming that the oral treatment of dogs with NexGard® prevented transmission of Babesia canis and development of clinical babesiosis for up to 28 days. Conclusion This is the first demonstration that an oral acaricidal treatment may prevent the transmission of a
Kamyingkird, Ketsarin; Cao, Shinuo; Tuvshintulga, Bumduuren; Salama, Akram; Mousa, Ahmed Abdelmoniem; Efstratiou, Artemis; Nishikawa, Yoshifumi; Yokoyama, Naoaki; Igarashi, Ikuo; Xuan, Xuenan
Theileria equi and Babesia caballi are the causative agents of equine piroplasmosis (EP), which affects equine production in various parts of the world. However, a safe and effective drug is not currently available for treatment of EP. Dihydroorotate dehydrogenase (DHODH) is the fourth enzyme in the de novo pyrimidine synthesis pathway and has been known as a novel drug target for several apicomplexan protozoan parasites. In this study, we evaluated four DHODH inhibitors; atovaquone (ATV), leflunomide (LFN), brequinar (Breq), and 7-hydroxy-5-[1,2,4] triazolo [1,5,a] pyrimidine (TAZ) on the growth of T. equi and B. caballi in vitro and compared them to diminacene aceturate (Di) as the control drug. The growth of T. equi and B. caballi was significantly hindered by all inhibitors except TAZ. The half maximal inhibitory concentration (IC50) of ATV, LFN, Breq and Di against T. equi was approximately 0.028, 109, 11 and 40 μM, respectively, whereas the IC50 of ATV, LFN, Breq and Di against B. caballi was approximately 0.128, 193, 5.2 and 16.2 μM, respectively. Using bioinformatics and Western blot analysis, we showed that TeDHODH was similar to other Babesia parasite DHODHs, and confirmed that targeting DHODHs could be useful for the development of novel chemotherapeutics for treatment of EP.
Petersen, Mette B; Krogh, Kaspar; Nielsen, Liza R
The relevance and limitations for using measurements of antibodies against Mycoplasma bovis in bulk tank milk (BTM) as a potentially cost-effective diagnostic tool for herd classification has not been evaluated before. Assuming that an increasing or high seroprevalence is a result of on-going or recent spread of M. bovis in a dairy herd, we tested the hypothesis that increasing prevalence of antibody-positive cows and young stock are associated with increasing BTM antibody ELISA values against M. bovis in Danish dairy herds with different courses of M. bovis infection. Furthermore, we tested whether herd size was associated with variations in the BTM responses. Thirty-nine Danish dairy herds selected to represent 4 different herd-level infection groups [8 control herds, 14 acute outbreak herds, 7 herds with previous outbreaks, and 10 herds with elevated BTM ELISA-values directed against M. bovis (>64% optical density measurement)] were visited 4 to 5 times, approximately 3mo apart. At each visit, 65 young stock were blood sampled. At the milk recording date closest to the herd visit date, 50 milk recording samples from individual lactating cows were randomly selected. In addition, a BTM sample was collected as a representative sample directly from the bulk tank by the dairies' milk truck drivers as part of the mandatory milk quality-control scheme. Blood and milk samples were tested for antibodies against M. bovis with a commercially available ELISA test (Bio-X BIO K 302, Bio-X Diagnostics, Rochefort, Belgium). A linear mixed effects model was used to analyze the effects of the prevalence of antibody-positive lactating cows and young stock and herd size on the BTM M. bovis ELISA results. Herd was included as a random effect to account for clustering of BTM samples originating from the same herd. Increasing prevalence of antibody-positive lactating cows was the only variable associated with increasing M. bovis BTM ELISA optical density measurement. In contrast, the
Jones, R M; Ashford, R; Cork, J; Palmer, S; Wood, E; Spyvee, P; Parks, S; Bennett, A; Brewer, J; Delahay, R; Chambers, M; Sawyer, J
Environmental air sampling was evaluated as a method to detect the presence of M. bovis in the vicinity of infected badgers and their setts. Airborne particles were collected on gelatine filters using a commercially available air sampling instrument and tested for the presence of M. bovis using bacteriological culture and real-time PCR. The sensitivity of bacteriological culture was broadly similar to that of real-time PCR when testing samples artificially spiked with M. bovis. Sampling was undertaken from directly under the muzzles of badgers which had been experimentally infected with M. bovis (37 samples), within enclosures housing the experimentally infected animals (50 samples), and in the vicinity of setts with resident infected wild badgers (52 samples). The methods employed did not detect M. bovis from either infected badgers or artificial or natural setts known to contain infected animals. However, samples taken at four of the six natural setts were positive for Mycobacterium gordonae.
Prieto, Claudia; Serra, Diego Omar; Martina, Pablo; Jacobs, Melisa; Bosch, Alejandra; Yantorno, Osvaldo M
The difficulties in preventing and treating infectious bovine keratoconjunctivitis (IBK) and the consequent impact on the cattle industry worldwide emphasize the need to better understand this infectious process along with the biology of Moraxella bovis, its primary causative agent. Although there is increasing evidence that bacterial biofilms participate in a variety of ocular infections by direct biofilm formation on the surfaces of the eye, IBK has not been considered as a biofilm-based disease so far, and even more, no information is currently available regarding the ability of M. bovis to adopt a biofilm lifestyle. In the present research, we demonstrated the capacity of M. bovis clinical isolates and reference strains to form biofilms on different abiotic surfaces and culture conditions, and provided qualitative and quantitative information on the biofilm growth and architecture of mature biofilms. In addition, our data indicated that the type IV pili play a critical role in the biofilm formation in vitro. Most significantly, we proved that through exposure to MgCl2 type IV pili are removed from the cell surface, not only preventing M. bovis biofilm formation but also disassembling preformed biofilms. These results could constitute a new approach in the understanding of M. bovis colonization process in cattle eye and/or nasal cavity, and may aid in the development of future antimicrobial strategies for the control of IBK.
KONG, Ling-Cong; GAO, Duo; JIA, Bo-Yan; WANG, Zi; GAO, Yun-Hang; PEI, Zhi-Hua; LIU, Shu-Ming; XIN, Jiu-Qing; MA, Hong-Xia
Mycoplasma bovis has spread widely throughout the world via animal movement and has become an important pathogen of bovine respiratory disease. However, the minimum inhibitory concentrations of antimicrobials for Mycoplasma bovis have not been studied in China. The objective of this study was to determine the prevalence and antibiotic resistance of Mycoplasma bovis isolated from young cattle with respiratory infection in China. Mycoplasma bovis was detected in 32/45 bovine respiratory infection outbreaks at beef farms in 8 provinces in China. The isolates were susceptible or had medium sensitivity to ciprofloxacin, enrofloxacin and doxycycline, but were frequently resistant to macrolides (13/32, 41%). An A2058G (Escherichia coli Numbering) mutation located in the rrnA operon in domain V of 23S rRNA was observed in strains that were resistant to macrolides. This single mutations at the rrnA operon in domain V of 23S rRNA may play an important role in the resistance of Mycoplasma bovis strains to macrolides. PMID:26346744
Rodríguez, Sabrina; Romero, Beatriz; Bezos, Javier; de Juan, Lucía; Alvarez, Julio; Castellanos, Elena; Moya, Nuria; Lozano, Francisco; González, Sergio; Sáez-Llorente, José Luis; Mateos, Ana; Domínguez, Lucas; Aranaz, Alicia
Mycobacterium bovis is the main causative agent of bovine tuberculosis. This zoonotic disease produces important economic losses and must be considered a threat to endangered animal species and public health. This study was performed (1) to assess the degree of diversity of the Spanish M. bovis isolates and its effect on the epidemiology of the infection, and (2) to understand the connection of M. bovis populations within a European context. In this report we resume the DVR-spoligotyping results of 6215 M. bovis isolates collected between 1992 and 2007 from different hosts. The isolates clustered into 252 spoligotypes which varied largely in frequency, geographical distribution and appearance in different animal species. In general, the most frequent spoligotypes were found all over the country and in different animal species, though some were restricted to a geographical area. Among our most often isolated spoligotypes, SB0121 and SB0120 (BCG-like) are a common feature between mainland European countries, however, the spoligotypes differ with those found in the UK, the Republic of Ireland and abroad. A comparison of spoligotypes reported from other countries reveals hints for the M. bovis demography in Europe and suggests a common ancestor strain. This study gives insight into the usefulness of the standardized DVR-spoligotyping technique for epidemiological studies in a country with a high degree of strain diversity.
Yisaschar-Mekuzas, Yael; Jaffe, Charles L; Pastor, Josep; Cardoso, Luís; Baneth, Gad
Canine infection by vector-borne hemoparasites is frequent in tropical and sub-tropical areas where exposure to hematophageous ectoparasites is intensive. A reverse line blot (RLB) assay was designed to improve the simultaneous detection of all named canine piroplasm species combined with other vector-borne pathogens of dogs including Ehrlichia canis, Hepatozoon canis and Leishmania infantum common in the Mediterranean basin. Blood samples of 110 dogs from Spain (n=21), Portugal (n=14) and Israel (n=75) were analyzed. The study evaluated 2 groups of dogs, 49 dogs with piroplasm infection detected by blood smear microscopy from Portugal, Spain and Israel, and 61 dogs surveyed from rural areas in Israel, for which infection status with vector-borne pathogens was unknown. Among the dogs previously diagnosed with piroplasmosis, infection with Babesia canis, Babesia vogeli, Babesia gibsoni and Theileria annae was detected in the Iberian dogs while only B. vogeli was found in Israeli dogs. These differences are attributed to the absence of tick vectors for some piroplasm species such as Dermacentor reticulatus in Israel. Eleven (79%) of the Babesia-positive dogs from Portugal were co-infected with other pathogens including L. infantum, H. canis and E. canis. Eight of 61 (13%) rural Israeli dogs were co-infected with two or more pathogens including B. vogeli, L. infantum, E. canis, and H. canis. Triple infections were demonstrated in 2 dogs. The RLB detection limit for Babesia was 50-fold lower than that of PCR. This study presents a RLB to simultaneously detect and separate the major vector-borne dog pathogens in southern Europe and the Middle East.
Takeet, Michael I; Oyewusi, Adeoye J; Abakpa, Simon A V; Daramola, Olukayode O; Peters, Sunday O
Adequate knowledge of the genetic diversity among Babesia species infecting dogs is necessary for a better understanding of the epidemiology and control of canine babesiosis. Hence, this study determined the genetic diversity among the Babesia rossi detected in dogs presented for routine examination in Veterinary Hospitals in Abeokuta, Nigeria. Blood were randomly collected from 209 dogs. Field-stained thin smears were made and DNA extracted from the blood. Partial region of the 18S small subunit ribosomal RNA (rRNA) gene was amplified, sequenced and analysed. Babesia species was detected in 16 (7.7%) of the dogs by microscopy. Electrophoresed PCR products from 39 (18.66%) dogs revealed band size of 450 bp and 2 (0.95%) dogs had band size of 430 bp. The sequences obtained from 450 bp amplicon displayed homology of 99.74% (387/388) with partial sequences of 18S rRNA gene of Babesia rossi in the GeneBank. Of the two sequences that had 430 bp amplicon, one was identified as T. annulata and second as T. ovis. A significantly (p<0.05) higher prevalence of B. rossi was detected by PCR compared to microscopy. The mean PCV of Babesia infected dogs was significantly (p<0.05) lower than non-infected dogs. Phylogenetic analysis revealed minimal diversity among B. rossi with the exception of one sequence that was greatly divergent from the others. This study suggests that more than one genotype of B. rossi may be in circulation among the dog population in the study area and this may have potential implication on clinical outcome of canine babesiosis.
Mycobacterium bovis is the cause of tuberculosis in most animal species, including cattle and is a serious zoonotic pathogen. In humans, M. bovis infection can result in disease clinically indistinguishable from that caused by Mycobacterium tuberculosis, the cause of most tuberculosis in humans. Reg...
Miller, R S; Sweeney, S J
Mycobacterium bovis (M. bovis), the causative agent of bovine tuberculosis, has been identified in nine geographically distinct wildlife populations in North America and Hawaii and is endemic in at least three populations, including members of the Bovidae, Cervidae, and Suidae families. The emergence of M. bovis in North American wildlife poses a serious and growing risk for livestock and human health and for the recreational hunting industry. Experience in many countries, including the USA and Canada, has shown that while M. bovis can be controlled when restricted to livestock species, it is almost impossible to eradicate once it has spread into ecosystems with free-ranging maintenance hosts. Therefore, preventing transmission of M. bovis to wildlife may be the most effective way to mitigate economic and health costs of this bacterial pathogen. Here we review the status of M. bovis infection in wildlife of North America and identify risks for its establishment in uninfected North American wildlife populations where eradication or control would be difficult and costly. We identified four common risk factors associated with establishment of M. bovis in uninfected wildlife populations in North America, (1) commingling of infected cattle with susceptible wildlife, (2) supplemental feeding of wildlife, (3) inadequate surveillance of at-risk wildlife, and (4) unrecognized emergence of alternate wildlife species as successful maintenance hosts. We then propose the use of integrated and adaptive disease management to mitigate these risk factors to prevent establishment of M. bovis in susceptible North American wildlife species.
Quirino, Angela; Torti, Carlo; Strazzulla, Alessio; Nisticò, Salvatore; Galati, Luisa; Barreca, Giorgio Settimo; Lamberti, Angelo Giuseppe; Berardelli, Giuseppina; Pacciarini, Maria; Gasparini, Giorgio; Pisani, Vincenzo; Gambardella, Antonio; Liberto, Maria Carla; Focà, Alfredo
We report herein the first case of a coinfection with Brucella spp., M. bovis, and Enterobacter cloacae in a butcher who moved from Bulgaria to Italy. Molecular typing suggested professional acquisition of M. bovis in Italy. So, surveillance and preventive measures need to be implemented. PMID:26257970
Efficacy of Oral and Parenteral Routes of Mycobacterium bovis bacille Calmette-Guerin Vaccination Against Experimental Bovine Tuberculosis in White-tailed deer (Odocoileus virginianus): A Feasibility Study
We investigated the efficacy of oral and parenteral Mycobacterium bovis bacille Calmette-Guerin Danish strain 1331 (BCG) in its ability to protect white-tailed deer (Odocoileus virginianus) against disease caused by M. bovis infection. Thirty white-tailed deer were divided into four groups. One gr...
Mundt, H-C; Bangoura, B; Mengel, H; Keidel, J; Daugschies, A
The efficacy of metaphylactic treatment with toltrazuril (Baycox 5% suspension) against natural infections with Eimeria bovis and/or Eimeria zuernii in calves was investigated. The study was conducted with 208 calves on five calf-rearing farms in Germany and the Czech Republic. All participating farms had a notable incidence of coccidiosis. Animals were treated 14 days after stabling in the respective facility. One group was treated with 15 mg toltrazuril/kg body weight, and a second group served as the sham-treated control. Assessment of efficacy was based on faecal consistency and oocyst excretion of E. bovis and E. zuernii, both investigated throughout the study. Duration and rate of oocyst excretion as well as number of scour days with E. bovis or E. zuernii oocyst shedding and the severity of diarrhoea were significantly lower in the toltrazuril-treated groups. It is concluded that a single metaphylactic treatment with toltrazuril controls coccidiosis of housed calves under various field conditions.
Miller, Michele; Buss, Peter; Hofmeyr, Jennifer; Olea-Popelka, Francisco; Parsons, Sven; van Helden, Paul
Diagnosis of tuberculosis in wildlife often relies on postmortem samples because of logistical challenges and lack of field-friendly techniques for live animal testing. Confirmation of infection through detection of infectious organisms is essential for studying the pathogenesis and epidemiology of disease. We describe the application of a technique to obtain respiratory samples from free-ranging living lions to facilitate detection of viable Mycobacterium bovis under field conditions. We identified M. bovis by mycobacterial culture and PCR in tracheobronchial lavage samples from 8/134 (6.0%) lions tested in Kruger National Park, South Africa. This confirms the respiratory shedding of viable M. bovis in living lions. The implications of these results are that infected lions have the potential to transmit this disease and serve as maintenance hosts.
Ma, Hongyue; Zhou, Jing; Jiang, Jiejun; Duan, Jinao; Xu, Huiqin; Tang, Yuping; Lv, Gaohong; Zhang, Junfeng; Zhan, Zhen; Ding, Anwei
Toad Venom, called chansu (CS) in China, is an anti-inflammatory drug used in small doses for the treatment of various types of inflammation in China. Its use is hampered by the cardiotoxicity of bufadienolides derived from Toad Venom. Bezoar Bovis is another frequently used drug in Toad Venom preparations for the treatment of inflammatory or cardiovascular diseases in Asia. We explored whether Bezoar Bovis could protect against CS-induced acute toxicity in mice. Toxicity was assessed by the general features of poisoning, electrocardiography (ECG), and levels of creatine kinase (CK), lactate dehydrogenase (LDH) and calcium ions (Ca(2+)) in cardiac tissues. Toad Venom (90 mg/kg) caused opisthotonus, ventricular arrhythmias, and increases in cardiac levels of Ca(2+), CK and LDH. Pretreatment with Bezoar Bovis (120, 240 and 480 mg/kg) significantly reduced the prevalence of opisthotonus and mortality, and prevented cardiotoxicity in CS-treated mice as evidenced by decreases in the scores of arrhythmias and cardiac levels of CK, LDH and Ca(2+). Furthermore, the bilirubin, and taurine derived from Bezoar Bovis offered marked protection against the arrhythmias induced by CS or bufalin in vivo and in vitro. An anti-inflammatory study showed that Bezoar Bovis did not compromise the anti-inflammatory activity of Toad Venom on concanavalin-A (ConA)-stimulated proliferation of human peripheral blood mononuclear cells. These results suggested that Bezoar Bovis elicited protective and anti-arrhythmic effects against Toad Venom intoxication in mice, and is a novel antidote in combination with Toad Venom therapy.
Moritz, Erin D; Winton, Colleen S; Tonnetti, Laura; Townsend, Rebecca L; Berardi, Victor P; Hewins, Mary-Ellen; Weeks, Karen E; Dodd, Roger Y; Stramer, Susan L
Background Babesia microti, a tickborne intraerythrocytic parasite that can be transmitted by means of blood transfusion, is responsible for the majority of cases of transfusion-transmitted babesiosis in the United States. However, no licensed test exists for screening for B. microti in donated blood. We assessed data from a large-scale, investigational product-release screening and donor follow-up program. Methods From June 2012 through September 2014, we performed arrayed fluorescence immunoassays (AFIAs) for B. microti antibodies and real-time polymerase-chain-reaction (PCR) assays for B. microti DNA on blood-donation samples obtained in Connecticut, Massachusetts, Minnesota, and Wisconsin. We determined parasite loads with the use of quantitative PCR testing and assessed infectivity by means of the inoculation of hamsters and the subsequent examination for parasitemia. Donors with test-reactive samples were followed. Using data on cases of transfusion-transmitted babesiosis, we compared the proportions of screened versus unscreened donations that were infectious. Results Of 89,153 blood-donation samples tested, 335 (0.38%) were confirmed to be positive, of which 67 (20%) were PCR-positive; 9 samples were antibody-negative (i.e., 1 antibody-negative sample per 9906 screened samples), representing 13% of all PCR-positive samples. PCR-positive samples were identified all through the year; antibody-negative infections occurred from June through September. Approximately one third of the red-cell samples from PCR-positive or high-titer AFIA-positive donations infected hamsters. Follow-up showed DNA clearance in 86% of the donors but antibody seroreversion in 8% after 1 year. In Connecticut and Massachusetts, no reported cases of transfusion-transmitted babesiosis were associated with screened donations (i.e., 0 cases per 75,331 screened donations), as compared with 14 cases per 253,031 unscreened donations (i.e., 1 case per 18,074 unscreened donations) (odds ratio, 8
Khatri, Bhagwati; Fielder, Mark; Jones, Gareth; Newell, William; Abu-Oun, Manal; Wheeler, Paul R.
Tuberculosis is a major human and animal disease of major importance worldwide. Genetically, the closely related strains within the Mycobacterium tuberculosis complex which cause disease are well-characterized but there is an urgent need better to understand their phenotypes. To search rapidly for metabolic differences, a working method using Biolog Phenotype MicroArray analysis was developed. Of 380 substrates surveyed, 71 permitted tetrazolium dye reduction, the readout over 7 days in the method. By looking for ≥5-fold differences in dye reduction, 12 substrates differentiated M. tuberculosis H37Rv and Mycobacterium bovis AF2122/97. H37Rv and a Beijing strain of M. tuberculosis could also be distinguished in this way, as could field strains of M. bovis; even pairs of strains within one spoligotype could be distinguished by 2 to 3 substrates. Cluster analysis gave three clear groups: H37Rv, Beijing, and all the M. bovis strains. The substrates used agreed well with prior knowledge, though an unexpected finding that AF2122/97 gave greater dye reduction than H37Rv with hexoses was investigated further, in culture flasks, revealing that hexoses and Tween 80 were synergistic for growth and used simultaneously rather than in a diauxic fashion. Potential new substrates for growth media were revealed, too, most promisingly N-acetyl glucosamine. Osmotic and pH arrays divided the mycobacteria into two groups with different salt tolerance, though in contrast to the substrate arrays the groups did not entirely correlate with taxonomic differences. More interestingly, these arrays suggested differences between the amines used by the M. tuberculosis complex and enteric bacteria in acid tolerance, with some hydrophobic amino acids being highly effective. In contrast, γ-aminobutyrate, used in the enteric bacteria, had no effect in the mycobacteria. This study proved principle that Phenotype MicroArrays can be used with slow-growing pathogenic mycobacteria and already has
Harrington, Noel P; Surujballi, Om P; Prescott, John F; Duncan, J Robert; Waters, W Ray; Lyashchenko, Konstantin; Greenwald, Rena
Captive and free-ranging wildlife animals are implicated in the maintenance and transmission of bovine tuberculosis and therefore pose a significant obstacle to eradication of the disease from domestic livestock. The current antemortem diagnostic method, the intradermal tuberculin skin test, is impractical for routine use with many wild animals. Antibody-based assays are particularly attractive because the animals are handled only once and immediate processing of the sample is not required. This report characterizes the antibody responses of red deer-elk hybrids (Cervus elaphus) against Mycobacterium bovis and subsequently evaluates the diagnostic performance of select antigens in a rapid-test format. Sequential serum samples were collected from 10 animals experimentally infected with M. bovis and 5 noninfected animals over a 7-month period postinfection (p.i.). Samples were evaluated by enzyme-linked immunosorbent assays, immunoblot analyses, and multiantigen print immunoassays for seroreactivity to mycobacterial antigens. Although all infected animals produced antibodies to M. bovis protein antigens, there was significant animal-to-animal variation in the kinetics and magnitudes of responses and the antigens recognized. The most frequently recognized antigens included MPB83, ESAT-6, CFP10, and MPB70. Responses to some antigens, such as MPB83, were consistently detected as early as 4 weeks after inoculation, whereas other antigens were detected only much later (>140 days p.i.). Antibody responses were boosted by injection of tuberculin for intradermal tuberculin skin testing. Comparison of single-antigen (fluorescence polarization assay) with multiantigen (CervidTB STAT-PAK) rapid tests demonstrated that a highly sensitive and specific serodiagnostic test for tuberculosis in cervids will require multiple and carefully selected seroreactive antigens covering a broad spectrum of antibody specificities.
Che' Amat, A; González-Barrio, D; Ortiz, J A; Díez-Delgado, I; Boadella, M; Barasona, J A; Bezos, J; Romero, B; Armenteros, J A; Lyashchenko, K P; Venteo, A; Rueda, P; Gortázar, C
Animal tuberculosis (TB) caused by infection with Mycobacterium bovis and closely related members of the M. tuberculosis complex (MTC), is often reported in the Eurasian wild boar (Sus scrofa). Tests detecting antibodies against MTC antigens are valuable tools for TB monitoring and control in suids. However, only limited knowledge exists on serology test performance in 2-6 month-old piglets. In this age-class, recent infections might cause lower antibody levels and lower test sensitivity. We examined 126 wild boar piglets from a TB-endemic site using 6 antibody detection tests in order to assess test performance. Bacterial culture (n=53) yielded a M. bovis infection prevalence of 33.9%, while serum antibody prevalence estimated by different tests ranged from 19% to 38%, reaching sensitivities between 15.4% and 46.2% for plate ELISAs and between 61.5% and 69.2% for rapid immunochromatographic tests based on dual path platform (DPP) technology. The Cohen kappa coefficient of agreement between DPP WTB (Wildlife TB) assay and culture results was moderate (0.45) and all other serological tests used had poor to fair agreements. This survey revealed the ability of several tests for detecting serum antibodies against the MTC antigens in 2-6 month-old naturally infected wild boar piglets. The best performance was demonstrated for DPP tests. The results confirmed our initial hypothesis of a lower sensitivity of serology for detecting M. bovis-infected piglets, as compared to older wild boar. Certain tests, notably the rapid animal-side tests, can contribute to TB control strategies by enabling the setup of test and cull schemes or improving pre-movement testing. However, sub-optimal test performance in piglets as compared to that in older wild boar should be taken into account.
Hamid, Penny H; Hirzmann, Jörg; Hermosilla, Carlos; Taubert, Anja
Eimeria bovis macromeront formation in bovine endothelial host cells is an energy- and nutrient-demanding process. Obligate intracellular replicating coccidians are generally considered as auxotrophic for cholesterol synthesis and scavenge cholesterol from the host cell by either enhancing the uptake of extracellular cholesterol sources or by upregulating the host cellular de novo biosynthesis. We here focused on the latter mechanism and analyzed the effects of several inhibitors targeting the host cellular mevalonate biosynthesis pathway and cholesterol processing. The following inhibitors were used: lovastatin, squalestatin, CI976 and C75 targeting HMG-CoA reductase, squalene synthase, acyl-CoA:cholesterol acyltransferase, and fatty acid synthase, respectively. In summary, all inhibitors significantly interfered with E. bovis meront formation and merozoite production in a dose-dependent manner. Dose effect responses identified lovastatin as the most effective compound, followed by CI976, C75, and squalestatin, respectively. Overall, merozoite production was inhibited by 99.6, 99.7, 84.6, and 70.2% via lovastatin (1 μM), CI976, C75, and squalestatin (all 5 μM) treatments, respectively. Concerning macromeront formation, both the rate and size of developing meronts were affected by inhibitor treatments. The effects were characterized by developmental arrest and meront degradation. In the case of CI976 treatment, we additionally observed detrimental effects on host cellular lipid droplet formation leading to meront developmental arrest irrespective of the time point of treatment onset. These analyses clearly indicate that successful E. bovis intracellular development strictly depends on the host cellular de novo biosynthesis of cholesterol and on the adequate subsequent processing thereof.
Background Bovine tuberculosis (BTB), caused by Mycobacterium bovis, has remained a major source of concern to public health officials in Zambia. Previous investigations have used traditional epidemiological methods that are unable to identify the causative agent and from which dynamics of disease dispersion is difficult to discern. The objective of this study was to isolate, characterize and determine the genetic diversity and relatedness of M. bovis from major cattle rearing districts in Zambia by spoligotyping. A total of 695 carcasses were examined and 98 tissues had gross post-mortem lesions compatible with BTB. Results Forty-two out of the ninety-eight suspected tissues examined had culture properties characteristic of mycobacteria from which 31 isolates yielded interpretable spoligotypes. This technique showed good discriminatory power (HGDI = 0.98), revealing 10 different spoligotype patterns. Twenty-seven isolates belonged to one cluster with more than 95% similarity and inside the cluster, one predominant spoligotype was found in 20 (64.5%) of the isolates tested. The highest number of spoligotypes was observed among samples from Namwala district. Spoligotypes from 26 (83.9%) of the isolates belonged to five spoligotypes that have been reported before while the remaining 5 (16.1%) isolates had unique spoligotypes that are being reported for the first time; these have been assigned numbers SB1763 to SB1767. Five of the 6 districts had the predominant spoligotype (SB0120). Conclusion The study has described the dispersion patterns of M. bovis in Zambian cattle for the first time and has identified 5 spoligotype patterns specific to Zambia. The observation of an overlap in the spoligotype pattern SB0120 in 5 of the 6 districts suggests the probability of sharing a common source of infection. PMID:19619309
Aagaard, Claus; Govaerts, Marc; Meng Okkels, Limei; Andersen, Peter; Pollock, John M.
Differential delayed-type hypersensitivity skin testing with tuberculin purified protein derivatives from Mycobacterium bovis and M. avium is the standard for diagnosing bovine tuberculosis. However, improved tests based on defined, specific antigens are urgently needed. In the present study, a combination of bioinformatics, molecular biology, and bovine models of infection were used to screen mycobacterial proteins for their potential as diagnostic reagents which could be used in a whole-blood assay for diagnosis of tuberculosis. Initial screening of 28 proteins selected in silico and expressed as recombinants in Escherichia coli indicated that CFP-10, ESAT-6, TB27.4, TB16.2, TB15.8, and TB10.4 induced strong gamma interferon responses in experimentally infected cattle. A more thorough investigation over time in two groups of animals infected with a high (106 CFU) and a low (104 CFU) dose of M. bovis revealed that, for both groups, the strength of the in vitro response to individual antigens varied greatly over time. However, combining the results for ESAT-6, CFP-10, and TB27.4, possibly supplemented with TB10.4, gave sensitivities at different infection stages close to those obtained with M. bovis purified protein derivative. Importantly, while responsiveness to ESAT-6 and CFP-10 correlated strongly for individual samples, the same was not the case for ESAT-6 and TB27.4 responsiveness. The results suggest that combinations of specific antigens such as these have great potential in development of optimized diagnostic systems for bovine tuberculosis. PMID:12904381
Ngendahayo, L D; Bayssade-Dufour, C; Albaret, J L; Diaw, O T; Deiana, S; Southgate, V R; Ross, G C; Luffau, G; Chabaud, A G
S. E. M. study of the dorsal anterior one third of male Schistosoma bovis and of the anterior ventral border of the gynaecophoric duct. S. bovis was previously described as possessing spineless tubercles. This is so in specimens obtained from experimentally infected rodents, but in cattle, on the contrary, when conditions are favourable, teguments have spiny tubercles. Two morphological types have been observed: the first in Bos taurus from Sardinia, the second in domestic (Bos indicus) and wild (Hippotragus equinus and Damaliscus korrigum) bovids from Senegal, Tchad and Centrafrican Republic.