Development of a shared item repository for progress testing in veterinary education.

Introduction: Progress testing in education is an assessment principle for the measurement of students' progress over time, e.g., from start to graduation. Progress testing offers valid longitudinal formative measurement of the growth in the cognitive skills of the individual students within the subjects of the test as well as a tool for educators to monitor potential educational gaps and mismatches within the curriculum in relation to the basic veterinary learning outcomes. Methods: Six veterinary educational establishments in Denmark, Finland, Germany (Hannover), the Netherlands, Norway, and Sweden established in cooperation with the European Association of Establishments for Veterinary Education (EAEVE) a common veterinary item repository that can be used for progress testing in European Veterinary Education Establishments (VEEs), linear as well as computer adaptive, covering the EAEVE veterinary subjects and theoretical "Day One Competencies." First, a blueprint was created, suitable item formats were identified, and a quality assurance process for reviewing and approving items was established. The items were trialed to create a database of validated and calibrated items, and the responses were subsequently psychometrically analyzed according to Modern Test Theory. Results: In total, 1,836 items were submitted of which 1,342 were approved by the reviewers for trial testing. 1,119 students from all study years and all partners VEEs participated in one or more of six item trials, and 1,948 responses were collected. Responses were analyzed using Rasch Modeling (analysis of item-fit, differential item function, item-response characteristics). A total of 821 calibrated items of various difficulty levels matching the veterinary students' abilities and covering the veterinary knowledge domains have been banked. Discussion: The item bank is now ready to be used for formative progress testing in European veterinary education. This paper presents and discusses possible pitfalls, problems, and solutions when establishing an international veterinary progress test.

Impacts of maternal microbiota and microbial metabolites on fetal intestine, brain, and placenta.

BACKGROUND: The maternal microbiota modulates fetal development, but the mechanisms of these earliest host-microbe interactions are unclear. To investigate the developmental impacts of maternal microbial metabolites, we compared full-term fetuses from germ-free and specific pathogen-free mouse dams by gene expression profiling and non-targeted metabolomics. RESULTS: In the fetal intestine, critical genes mediating host-microbe interactions, innate immunity, and epithelial barrier were differentially expressed. Interferon and inflammatory signaling genes were downregulated in the intestines and brains of the fetuses from germ-free dams. The expression of genes related to neural system development and function, translation and RNA metabolism, and regulation of energy metabolism were significantly affected. The gene coding for the insulin-degrading enzyme (Ide) was most significantly downregulated in all tissues. In the placenta, genes coding for prolactin and other essential regulators of pregnancy were downregulated in germ-free dams. These impacts on gene expression were strongly associated with microbially modulated metabolite concentrations in the fetal tissues. Aryl sulfates and other aryl hydrocarbon receptor ligands, the trimethylated compounds TMAO and 5-AVAB, Glu-Trp and other dipeptides, fatty acid derivatives, and the tRNA nucleobase queuine were among the compounds strongly associated with gene expression differences. A sex difference was observed in the fetal responses to maternal microbial status: more genes were differentially regulated in male fetuses than in females. CONCLUSIONS: The maternal microbiota has a major impact on the developing fetus, with male fetuses potentially more susceptible to microbial modulation. The expression of genes important for the immune system, neurophysiology, translation, and energy metabolism are strongly affected by the maternal microbial status already before birth. These impacts are associated with microbially modulated metabolites. We identified several microbial metabolites which have not been previously observed in this context. Many of the potentially important metabolites remain to be identified.

Faecal microbiota in two-week-old female dairy calves during acute cryptosporidiosis outbreak - Association with systemic inflammatory response.

In the present study, relationships between the intestinal microbiota and innate immunity response, acute cryptosporidiosis, and weight gain in female dairy calves were investigated. A total of 112 calves born during a natural outbreak of cryptosporidiosis on one dairy farm was included in the study. Microbiota composition was analysed by means of 16S ribosomal RNA gene amplicon sequencing from faecal samples collected during the second week of life, while the status of Cryptosporidium spp. infection was determined using immunofluorescence. Serum samples from the second week of life were colourimetrically analysed for the following markers of acute inflammation: acute-phase proteins (serum amyloid A and haptoglobin) and pro-inflammatory cytokines (interleukin-1 beta, interleukin-6, and tumour necrosis factor-alpha). Statistical analyses were performed using random forest analysis, variance-partitioning, and negative binomial regression. The faecal microbiota of the two-week old calves was composed of the phyla Firmicutes, Bacteroidetes, Proteobacteria, Fusobacteria, and Actinobacteria (in order of decreasing abundance). Microbial diversity, measured in terms of the Shannon index, increased with the age of the calves and decreased if a high count of Cryptosporidium spp. oocysts was found in the faeces. Fusobacterium was positively associated with Cryptosporidium spp. oocyst count and serum amyloid A concentration. Peptostreptococcus was positively associated with haptoglobin and serum amyloid A concentrations, and negatively associated with average daily weight gain at 9 months of age. The markers of innate immunity, in combination with age, explained 6% of the microbial variation. These results suggest that some components of the intestinal microbiota may have a long-lasting negative effect on animal growth through the stimulation of the systemic innate immune response.

Maternal microbiota-derived metabolic profile in fetal murine intestine, brain and placenta.

BACKGROUND: The maternal microbiota affects the development of the offspring by microbial metabolites translocating to the fetus. To reveal the spectrum of these molecular mediators of the earliest host-microbe interactions, we compared placenta, fetal intestine and brain from germ-free (GF) and specific pathogen free (SPF) mouse dams by non-targeted metabolic profiling. RESULTS: One hundred one annotated metabolites and altogether 3680 molecular features were present in significantly different amounts in the placenta and/or fetal organs of GF and SPF mice. More than half of these were more abundant in the SPF organs, suggesting their microbial origin or a metabolic response of the host to the presence of microbes. The clearest separation was observed in the placenta, but most of the molecular features showed significantly different levels also in the fetal intestine and/or brain. Metabolites that were detected in lower amounts in the GF fetal organs included 5-aminovaleric acid betaine, trimethylamine N-oxide, catechol-O-sulphate, hippuric and pipecolic acid. Derivatives of the amino acid tryptophan, such as kynurenine, 3-indolepropionic acid and hydroxyindoleacetic acid, were also less abundant in the absence of microbiota. Ninety-nine molecular features were detected only in the SPF mice. We also observed several molecular features which were more abundant in the GF mice, possibly representing precursors of microbial metabolites or indicators of a metabolic response to the absence of microbiota. CONCLUSIONS: The maternal microbiota has a profound impact on the fetal metabolome. Our observations suggest the existence of a multitude of yet unidentified microbially modified metabolites which pass through the placenta into the fetus and potentially influence fetal development.

The Composition of the Microbiota in the Full-Term Fetal Gut and Amniotic Fluid: A Bovine Cesarean Section Study.

The development of a healthy intestinal immune system requires early microbial exposure. However, it remains unclear whether microbial exposure already begins at the prenatal stage. Analysis of such low microbial biomass environments are challenging due to contamination issues. The aims of the current study were to assess the bacterial load and characterize the bacterial composition of the amniotic fluid and meconium of full-term calves, leading to a better knowledge of prenatal bacterial seeding of the fetal intestine. Amniotic fluid and rectal meconium samples were collected during and immediately after elective cesarean section, performed in 25 Belgian Blue cow-calf couples. The samples were analyzed by qPCR, bacterial culture using GAM agar and 16S rRNA gene amplicon sequencing. To minimize the effects of contaminants, we included multiple technical controls and stringently filtered the 16S rRNA gene sequencing data to exclude putative contaminant sequences. The meconium samples contained a significantly higher amount of bacterial DNA than the negative controls and 5 of 24 samples contained culturable bacteria. In the amniotic fluid, the amount of bacterial DNA was not significantly different from the negative controls and all samples were culture negative. Bacterial sequences were identified in both sample types and were primarily of phyla Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria, with some individual variation. We conclude that most calves encounter in utero maternal-fetal transmission of bacterial DNA, but the amount of bacterial DNA is low and viable bacteria are rare.

An across-breed validation study of 46 genetic markers in canine hip dysplasia.

BACKGROUND: Canine hip dysplasia (CHD) is a common disease, with a complex genetic background. Dogs with severe CHD sometimes also suffer from osteoarthritis (OA), an inflammatory, often painful and incurable condition. Previous studies have reported breed-specific genetic loci associated with different hip dysplasia and OA phenotypes. However, the independent replication of the known associations within or across breeds has been difficult due to variable phenotype measures, inadequate sample sizes and the existence of population specific variants. RESULTS: We execute a validation study of 46 genetic markers in a cohort of nearly 1600 dogs from ten different breeds. We categorize the dogs into cases and controls according to the hip scoring system defined by the Fédération Cynologique Internationale (FCI). We validate 21 different loci associated on fourteen chromosomes. Twenty of these associated with CHD in specific breeds, whereas one locus is unique to the across-breed study. We show that genes involved in the neddylation pathway are enriched among the genes in the validated loci. Neddylation contributes to many cellular functions including inflammation. CONCLUSIONS: Our study successfully replicates many loci and highlights the complex genetic architecture of CHD. Further characterisation of the associated loci could reveal CHD-relevant genes and pathways for improved understanding of the disease pathogenesis.

Genetic dissection of canine hip dysplasia phenotypes and osteoarthritis reveals three novel loci.

BACKGROUND: Hip dysplasia and osteoarthritis continue to be prevalent problems in veterinary and human medicine. Canine hip dysplasia is particularly problematic as it massively affects several large-sized breeds and can cause a severe impairment of the quality of life. In Finland, the complex condition is categorized to five classes from normal to severe dysplasia, but the categorization includes several sub-traits: congruity of the joint, Norberg angle, subluxation degree of the joint, shape and depth of the acetabulum, and osteoarthritis. Hip dysplasia and osteoarthritis have been proposed to have separate genetic etiologies. RESULTS: Using Fédération Cynologique Internationale -standardized ventrodorsal radiographs, German shepherds were rigorously phenotyped for osteoarthritis, and for joint incongruity by Norberg angle and femoral head center position in relation to dorsal acetabular edge. The affected dogs were categorized into mild, moderate and severe dysplastic phenotypes using official hip scores. Three different genome-wide significant loci were uncovered. The strongest candidate genes for hip joint incongruity were noggin (NOG), a bone and joint developmental gene on chromosome 9, and nanos C2HC-type zinc finger 1 (NANOS1), a regulator of matrix metalloproteinase 14 (MMP14) on chromosome 28. Osteoarthritis mapped to a long intergenic region on chromosome 1, between genes encoding for NADPH oxidase 3 (NOX3), an intriguing candidate for articular cartilage degradation, and AT-rich interactive domain 1B (ARID1B) that has been previously linked to joint laxity. CONCLUSIONS: Our findings highlight the complexity of canine hip dysplasia phenotypes. In particular, the results of this study point to the potential involvement of specific and partially distinct loci and genes or pathways in the development of incongruity, mild dysplasia, moderate-to-severe dysplasia and osteoarthritis of canine hip joints. Further studies should unravel the unique and common mechanisms for the various sub-traits.

Novel protective and risk loci in hip dysplasia in German Shepherds.

Canine hip dysplasia is a common, non-congenital, complex and hereditary disorder. It can inflict severe pain via secondary osteoarthritis and lead to euthanasia. An analogous disorder exists in humans. The genetic background of hip dysplasia in both species has remained ambiguous despite rigorous studies. We aimed to investigate the genetic causes of this disorder in one of the high-risk breeds, the German Shepherd. We performed genetic analyses with carefully phenotyped case-control cohorts comprising 525 German Shepherds. In our genome-wide association studies we identified four suggestive loci on chromosomes 1 and 9. Targeted resequencing of the two loci on chromosome 9 from 24 affected and 24 control German Shepherds revealed deletions of variable sizes in a putative enhancer element of the NOG gene. NOG encodes for noggin, a well-described bone morphogenetic protein inhibitor affecting multiple developmental processes, including joint development. The deletion was associated with the healthy controls and mildly dysplastic dogs suggesting a protective role against canine hip dysplasia. Two enhancer variants displayed a decreased activity in a dual luciferase reporter assay. Our study identifies novel loci and candidate genes for canine hip dysplasia, with potential regulatory variants in the NOG gene. Further research is warranted to elucidate how the identified variants affect the expression of noggin in canine hips, and what the potential effects of the other identified loci are.

Publisher Correction: The composition of the perinatal intestinal microbiota in cattle.

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The composition of the perinatal intestinal microbiota in cattle.

Recent research suggests that the microbial colonization of the mammalian intestine may begin before birth, but the observations are controversial due to challenges in the reliable sampling and analysis of low-abundance microbiota. We studied the perinatal microbiota of calves by sampling them immediately at birth and during the first postnatal week. The large size of the bovine newborns allows sampling directly from rectum using contamination-shielded swabs. Our 16S rDNA data, purged of potential contaminant sequences shared with negative controls, indicates the existence of a diverse low-abundance microbiota in the newborn rectal meconium and mucosa. The newborn rectal microbiota was composed of Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes. The microbial profile resembled dam oral rather than fecal or vaginal vestibular microbiota, but included typical intestinal taxa. During the first postnatal day, the rectum was invaded by Escherichia/Shigella and Clostridia, and the diversity collapsed. By 7 days, diversity was again increasing. In terms of relative abundance, Proteobacteria were replaced by Firmicutes, Bacteroidetes and Actinobacteria, including Faecalibacterium, Bacteroides, Lactobacillus, Butyricicoccus and Bifidobacterium. Our observations suggest that mammals are seeded before birth with a diverse microbiota, but the microbiota changes rapidly in the early postnatal life.

Altering a computerized laboratory test order form rationalizes ordering of laboratory tests in primary care physicians.

BACKGROUND: To reduce physicians' inappropriate laboratory requests for their patients, administrators have used methods such as modifying a laboratory request order form with an agreed requesting protocol for the most common diagnoses in primary health care. OBJECTIVE: To study the effects of removing the erythrocyte sedimentation rate (ESR) and aspartate transaminase (AST) which are considered of limited clinical value for primary care clinical decision-making from a computerized laboratory test order form. These tests were removed to another new view from the electronic laboratory menu where the physicians, instead of just ticking the desired test from the list, had to do 4-8s extra work by writing down the abbreviation to order the test. METHODS: An observational controlled prospective study based on a before-after design was performed by removing AST and ES from the laboratory test order form of the computerized laboratory system for all primary care in the city of Helsinki, Finland. The numbers of annual and monthly use of AST and ESR and their controls, alanine transaminase (ALT) and C-reactive protein (CRP) ordered by General practitioners (GPs) was recorded over an eight-year period: four years before and a four years after the removal of AST and ES. RESULTS: Removing AST and ESR from the computerized laboratory test order form decreased their use by up to 90%, whereas the use of the control tests increased throughout the follow-up period. The variation in use of these removed tests also decreased. CONCLUSION: Removing a laboratory test from a computerized laboratory test order form may significantly reduce GPs' use of the laboratory test. Further studies are needed, however, to ensure the safety of this type of intervention.

Teat Morphology Characterization With 3D Imaging.

The objective of this study was to visualize, in a novel way, the morphological characteristics of bovine teats to gain a better understanding of the detailed teat morphology. We applied silicone casting and 3D digital imaging in order to obtain a more detailed image of the teat structures than that seen in previous studies. Teat samples from 65 dairy cows over 12 months of age were obtained from cows slaughtered at an abattoir. The teats were classified according to the teat condition scoring used in Finland and the lengths of the teat canals were measured. Silicone molds were made from the external teat surface surrounding the teat orifice and from the internal surface of the teat consisting of the papillary duct, Fürstenberg's rosette, and distal part of the teat cistern. The external and internal surface molds of 35 cows were scanned with a 3D laser scanner. The molds and the digital 3D models were used to evaluate internal and external teat surface morphology. A number of measurements were taken from the silicone molds. The 3D models reproduced the morphology of the teats accurately with high repeatability. Breed didn't correlate with the teat classification score. The rosette was found to have significant variation in its size and number of mucosal folds. The internal surface morphology of the rosette did not correlate with the external surface morphology of the teat implying that it is relatively independent of milking parameters that may impact the teat canal and the external surface of the teat.

Genomics and Proteomics Provide New Insight into the Commensal and Pathogenic Lifestyles of Bovine- and Human-Associated Staphylococcus epidermidis Strains.

The present study reports comparative genomics and proteomics of Staphylococcus epidermidis (SE) strains isolated from bovine intramammary infection (PM221) and human hosts (ATCC12228 and RP62A). Genome-level profiling and protein expression analyses revealed that the bovine strain and the mildly infectious ATCC12228 strain are highly similar. Their genomes share high sequence identity and synteny, and both were predicted to encode the commensal-associated fdr marker gene. In contrast, PM221 was judged to differ from the sepsis-associated virulent human RP62A strain on the basis of distinct protein expression patterns and overall lack of genome synteny. The 2D DIGE and phenotypic analyses suggest that PM221 and ATCC12228 coordinate the TCA cycle activity and the formation of small colony variants in a way that could result in increased viability. Pilot experimental infection studies indicated that although ATCC12228 was able to infect a bovine host, the PM221 strain caused more severe clinical signs. Further investigation revealed strain- and condition-specific differences among surface bound proteins with likely roles in adhesion, biofilm formation, and immunomodulatory functions. Thus, our findings revealed a close link between the bovine and commensal-type human strains and suggest that humans could act as a reservoir of bovine mastitis-causing SE strains.

Comparative proteome profiling of bovine and human Staphylococcus epidermidis strains for screening specifically expressed virulence and adaptation proteins.

The present study reports a comparative proteome cataloging of a bovine mastitis and a human-associated Staphylococcus epidermidis strain with a specific focus on surfome (cell-wall bound and extracellular) proteins. Protein identification by 1DE coupled with LC-MS/MS analyses resulted in 1400 and 1287 proteins from the bovine (PM221) and human (ATCC12228) strains, respectively, covering over 50% of all predicted and more than 30% of all predicted surfome proteins in both strains. Comparison of the identification results suggests elevated levels of proteins involved in adherence, biofilm formation, signal transduction, house-keeping functions, and immune evasion in PM221, whereas ATCC12228 was more effective in expressing host defense evasion proteases, skin adaptation lipases, hemagglutination, and heavy-metal resistance proteins. Phenotypic analyses showed that only PM221 displays protein- and DNA-mediated adherent growth, and that PM221 was more efficient in cleaving tributyrin, a natural compound of milk fat under low CO2 conditions. These findings are in line with the identification data and suggest that distinct expression of lipases and adhesive surfome proteins could lead to the observed phenotypes. This study is the first extensive survey of S. epidermidis proteomes to date, providing several protein candidates to be examined for their roles in adaptation and virulence in vivo. All MS data have been deposited in the ProteomeXchange with identifier PXD000404 (http://proteomecentral.proteomexchange.org/dataset/PXD000404).

Expansion of the preimmune antibody repertoire by junctional diversity in Bos taurus.

Cattle have a limited range of immunoglobulin genes which are further diversified by antigen independent somatic hypermutation in fetuses. Junctional diversity generated during somatic recombination contributes to antibody diversity but its relative significance has not been comprehensively studied. We have investigated the importance of terminal deoxynucleotidyl transferase (TdT) -mediated junctional diversity to the bovine immunoglobulin repertoire. We also searched for new bovine heavy chain diversity (IGHD) genes as the information of the germline sequences is essential to define the junctional boundaries between gene segments. New heavy chain variable genes (IGHV) were explored to address the gene usage in the fetal recombinations. Our bioinformatics search revealed five new IGHD genes, which included the longest IGHD reported so far, 154 bp. By genomic sequencing we found 26 new IGHV sequences that represent potentially new IGHV genes or allelic variants. Sequence analysis of immunoglobulin heavy chain cDNA libraries of fetal bone marrow, ileum and spleen showed 0 to 36 nontemplated N-nucleotide additions between variable, diversity and joining genes. A maximum of 8 N nucleotides were also identified in the light chains. The junctional base profile was biased towards A and T nucleotide additions (64% in heavy chain VD, 52% in heavy chain DJ and 61% in light chain VJ junctions) in contrast to the high G/C content which is usually observed in mice. Sequence analysis also revealed extensive exonuclease activity, providing additional diversity. B-lymphocyte specific TdT expression was detected in bovine fetal bone marrow by reverse transcription-qPCR and immunofluorescence. These results suggest that TdT-mediated junctional diversity and exonuclease activity contribute significantly to the size of the cattle preimmune antibody repertoire already in the fetal period.

Mycobacterium marinum causes a latent infection that can be reactivated by gamma irradiation in adult zebrafish.

The mechanisms leading to latency and reactivation of human tuberculosis are still unclear, mainly due to the lack of standardized animal models for latent mycobacterial infection. In this longitudinal study of the progression of a mycobacterial disease in adult zebrafish, we show that an experimental intraperitoneal infection with a low dose (≈ 35 bacteria) of Mycobacterium marinum, results in the development of a latent disease in most individuals. The infection is characterized by limited mortality (25%), stable bacterial loads 4 weeks following infection and constant numbers of highly organized granulomas in few target organs. The majority of bacteria are dormant during a latent mycobacterial infection in zebrafish, and can be activated by resuscitation promoting factor ex vivo. In 5-10% of tuberculosis cases in humans, the disease is reactivated usually as a consequence of immune suppression. In our model, we are able to show that reactivation can be efficiently induced in infected zebrafish by γ-irradiation that transiently depletes granulo/monocyte and lymphocyte pools, as determined by flow cytometry. This immunosuppression causes reactivation of the dormant mycobacterial population and a rapid outgrowth of bacteria, leading to 88% mortality in four weeks. In this study, the adult zebrafish presents itself as a unique non-mammalian vertebrate model for studying the development of latency, regulation of mycobacterial dormancy, as well as reactivation of latent or subclinical tuberculosis. The possibilities for screening for host and pathogen factors affecting the disease progression, and identifying novel therapeutic agents and vaccine targets make this established model especially attractive.

The bovine genomic DNA sequence data reveal three IGHV subgroups, only one of which is functionally expressed.

A comprehensive analysis of cattle shotgun sequencing data reveals 36 immunoglobulin heavy chain variable genes. The previously described bovine subgroup IGHV1 contains 10 functional genes with a conserved promoter including the consensus octamer and several other transcription factor binding sites, intact exons and matching cDNA sequences. Subgroups IGHV2 and IGHV3 consist entirely of pseudogenes. Thus, the bovine germline IGHV repertoire is very limited. The IGHV genes are distributed in mammalian clans I and II, while no clan III genes were detected. Clan-specific PCR of genomic DNA from cattle, sheep, Eurasian elk, white-tailed deer, pig and dolphin indicates highly dynamic evolution of IGHV gene usage within Cetartiodactyla. The bovine germline IGHV repertoire was probably generated by recent duplications of an IGHV1-IGHV2 homology unit. Immunoglobulin heavy chain genes are largely incorrectly assembled in the current cattle genome versions Btau_4.2 and UMD_3.1. FISH experiments confirm an IGHV locus close to terminus of BTA21.

B lymphopoiesis is characterized by pre-B cell marker gene expression in fetal cattle and declines in adults.

Fetal cattle B-cell development proceeds via a pre-B cell stage that is characterized by the expression of surrogate light chain and recombination activation genes. In this paper, we identify a new member of bovine pre-B lymphocyte genes, VPREB2. Using RT-qPCR, we assess the expression of VPREB2 and three other surrogate light chain genes as well as RAG1 and RAG2 in fetal and adult cattle tissues. The absence of VPREB1, IGLL1, RAG1 and RAG2 expression in adult tissues and the lack of B-lymphoid differentiation in adult bone marrow - OP9 stromal cell co-culture, suggest a decline of B lymphopoiesis in adult cattle. The marked differences in the expression profiles of VPREB2 and VPREB3 in comparison to those of VPREB1, IGLL1 and RAGs suggest that the biological roles of VPREB2 and VPREB3 are unrelated to the pre-B cells.

Fetal bovine bone marrow is a rich source of CD34+ hematopoietic progenitors with myelo-monocytic colony-forming activity.

The CD34 glycoprotein is an important marker of hematopoietic stem cells. We used a polyclonal rabbit anti-bovine CD34 antibody to stain fetal and adult bovine bone marrow cells. Flow cytometry revealed a low side scatter (SSC(low)) population of cells that were CD34(+) but negative for leukocyte lineage markers CD11b, CD14 or CD2. Hematopoietic colony assays with CD34(+) and CD34(-) bone marrow cells suggested that the colony-forming potential in SSC(low) bone marrow cells was confined to the CD34(+) fraction. In contrast, this population was not enriched for cells expressing high aldehyde dehydrogenase activity, a metabolic marker that has been used to characterize hematopoietic stem cells. Thus, the CD34 antigen can be used to identify and isolate bovine bone marrow cells exhibiting clonogenic potential in vitro. Moreover, the proportion of CD34(+) cells is very high in fetal bovine bone marrow, indicating it as a rich source of hematopoietic progenitors.