An in vitro study of hematopoietic progenitor cells from peripheral blood of rabbits

The purpose of this study was to isolate and cultivate a subpopulation of pluripotent stem cells (PSCs) from the peripheral blood of rabbits, which are frequently used in veterinary research as an animal model. Pluripotent stem cells, as described in human beings, are fibroblast-like cells that exhibit a CD34 marker, specific from other hematopoietic stem cells. Commonly used human commercial media has been researched for culturing rabbit PSCs. These findings allow us to contemplate the direct application of this simple and standardized methodology in several areas of study, such as of the pharmacological effect of many drugs on hematopoietic cells, veterinary practice, and even the study of new strategies in cellular therapy for some human diseases.     

Study and culture of haematopoietic progenitor cells from peripheral blood in rats, hamsters and mice

The aim of this work was to isolate and cultivate a subpopulation of pluripotent stem cells present in peripheral blood of different animal species, frequently used in laboratory studies (mice, rats and hamsters). Pluripotent stem cells (PSCs), already described in human beings, are fibroblast-like cells that exhibit a CD34 marker, specific for haematopoietic stem cells. Commonly used human commercial media were investigated for culturing animal PSCs. These findings suggest that this simple and standardized methodology may be applicable in several fields such as the study of the pharmacological effects of drugs on the haematopoietic line and the study of new strategies in cellular therapy for some human diseases.     

Epizootiology and vectors of African horse sickness virus

African horse sickness (AHS) virus causes a non-contagious, infectious, arthropod-borne disease of equines and is enzootic in sub-Saharan Africa. The major vectors are species of Culicoides but mosquitoes and ticks may be involved. Periodically the virus makes excurisons beyond its enzootic zones but until recently has not been able to maintain itself outside these areas for more than 2–3 consecutive years. This is probably due to a number of factors including the absence of a long term vertebrate reservoir, the prevalence and seasonal incidence of the vectors and the efficiency of control measures. The recent AHS epizootics in Iberia and North Africa seem to have established a new pattern in AHS virus persistence. This is probably linked to the continuous presence of adult C. imicola in the area. Culicoides imicola is basically an Afro-Asiatic insect and prefers warm climates. Therefore its continuous adult presence in parts of Iberia may be due to some recent moderation of the climate in these areas.     

Comparison of a commercialized phenotyping system, antimicrobial susceptibility testing, and tuf gene sequence-based genotyping for species-level identification of coagulase-negative staphylococci isolated from cases of bovine mastitis

In order to evaluate the usefulness of some phenotypic and genotypic methods for species identification of coagulase-negative staphylococci (CNS), isolates were obtained from bovine cases of clinical and sub-clinical mastitis from different geographical areas in Sweden. By using the Staph-Zym test, antimicrobial susceptibility testing, and sequencing of part of the CNS tuf gene and, when needed, part of the 16S rRNA gene we characterized 82 clinical isolates and 24 reference strains of 18 different species of staphylococci. The genotypic methods identified nine different species of CNS among the 82 milk isolates. A comparison with results obtained by tuf gene sequencing showed that Staph-Zym correctly identified CNS reference strains to species level more often than bovine milk CNS isolates (83% and 61%, respectively). In addition, tests supplementary to the Staph-Zym were frequently needed in both groups of isolates (50% of reference strains and 33% of milk isolates) to obtain an identification of the strain. It is notable that Staph-Zym judged two isolates as CNS, although they belonged to other species, could not give a species name in 11% of the bovine CNS isolates, and gave 28% of the isolates an incorrect species name. The present study indicates that the studied phenotypic methods are unreliable for identification of CNS from bovine intra-mammary infections.     

Filling the gaps in the Argentinian distribution of orthohantavirus: First finding of Lechiguanas virus in rodents from Corrientes,Argentina

Hantavirus Pulmonary Syndrome (HPS) is an emerging infectious disease caused by orthohantaviruses in the Americas. In Argentina, since 1995, several reservoirs and virus variants have been described, but the northeastern and central endemic zones in the country include an area without human or rodent infections, despite sharing rodent species with areas with that disease. The aim of this study was to search for orthohantavirus in rodent communities that inhabit this area, which borders two endemic areas of HPS. Small rodents were captured in June of 2022 through a total effort of 644 trap nights distributed in five grids located in the Iberá National Park, Corrientes, Northeastern Argentina. All rodents were sexed, weighed, and the species was recorded. Blood samples were extracted to detect ANDV-specific immunoglobulin G (IgG), and to extract the RNA virus. Trimmed sequences were mapped against reference sequences from GenBank. We captured a total of 36 Oligoryzomys flavescens and 15 Oxymycterus rufus. We detected the O. flavescens species infected with Lechiguanas orthohantavirus in the camping area of the National Park. A nucleotide comparison with previously published sequences shows a 98.34% similarity to the virus obtained from a human case of HPS reported in the adjacent Misiones province. This study demonstrated, for the first time, that O. flavescens is a host of the Lechiguanas orthohantavirus in this zone and contributes to closing information gaps on the distribution of orthohantavirus in Argentina. Additionally, the high similarity with the hantavirus found in the human case of Misiones suggests that the reservoir in that province would also be O. flavescens (not previously confirmed). This information permits us to focus on the preventive measurements to protect the human population.     

北京油鸡角膜缘干细胞分离培养及分化潜能研究

【目的】 研究北京油鸡角膜缘干细胞（LSCs）分离、培养、鉴定及多向分化潜能等生物学特性。【方法】 取10胚龄健康北京油鸡鸡胚的角膜缘,采用分散酶Ⅱ（dispaseⅡ）联合胰蛋白酶二步法分离细胞,进行体外培养,绘制生长曲线,通过免疫荧光和RT-PCR技术鉴定LSCs特异性标志物细胞角蛋白3（CK3）、CK12、细胞周期调控蛋白p63、ATP结合盒转运蛋白（ABCG2）和细胞角蛋白19（CK19）的表达;通过成骨和成软骨诱导检测其多向分化潜能。【结果】 获得的北京油鸡细胞折光性强、贴壁后呈多角形,生长曲线呈典型的S型,群体倍增时间（PDT）随着代次的升高逐渐下降;免疫荧光检测结果表明,分离的细胞中ABCG2、p63和CK19均呈阳性表达,CK3和CK12不表达;RT-PCR检测结果表明,分离的细胞表达p63、CK19、ABCG2,不表达CD45,说明分离的细胞为LSCs。LCSs成骨诱导分化后,可见被茜素红染色的矿化钙结节,RT-PCR检测结果表明其表达成骨关键基因骨桥蛋白（OPN）和Ⅰ型胶原蛋白（Col-Ⅰ）;成软骨诱导分化后,可见被阿利新蓝染色的软骨组织,RT-PCR检测结果表明其表达成软骨细胞关键基因转录因子性别决定区Y框9（SOX-9）和Ⅱ型胶原蛋白（Col-Ⅱ）。【结论】 本研究成功从10胚龄北京油鸡胚胎角膜缘中分离LSCs,且其具有良好的增殖活力及成骨和成软骨能力,结果可为禽类LSCs的资源保存研究提供参考。     

Isolation and development of haematopoietic progenitor cells from peripheral blood of adult and newborn pigs

Pluripotent stem cells (PSCs), already described in human beings, are fibroblast-like cells that exhibit a CD34 marker specific for haematopoietic stem cells. In this work we have demonstrated the presence of PSCs in the peripheral blood of pigs, a species frequently used in transplantation studies as an animal model for human diseases. Differentiation into haematopoietic colonies (granulomacrophagic colonies, erythroid colonies and mixed colonies) has been carried out with the peripheral blood of adult and newborn pigs, using solely human commercial media. Peripheral blood mononuclear cells (PBMNCs) were cultured in semisolid methylcellulose based media enriched with recombinant human cytokines, achieving granulomacrophagic-colony forming unit (GM-CFU) and mixed-colony forming unit (Mix-CFU) growth with erythroblastic lineage proliferation in the presence of erythropoietin (Epo). In all the samples CFU growth was associated with the presence of recombinant human cytokine. No evidence of proliferation in control plates without cytokines was found. From liquid medium culture, a population of macrophages and CD34+ fibroblast like cells were retrieved 21 days after sowing. These findings allow us to think about the direct application of this simple and standardised method in several work fields such as the study of pharmacological effects of many drugs over the haematopoietic line and in the study of new strategies in cellular therapy for some human diseases.     

Isolation,proliferation and characterization of endometrial canine stem cells

In the last decade, progenitor cells isolated from dissociated endometrial tissue have been the subject of many studies in several animal species. Recently, endometrial cells showing characteristics of mesenchymal stem cells (MSC) have been demonstrated in human, pig and cow uterine tissue samples. The aim of this study was the isolation and characterization of stromal cells from the endometrium of healthy bitches, a tissue that after elective surgery is routinely discarded. Multipotent stromal cells could be isolated from all bitches enrolled in the study (n = 7). The multipotency of cells was demonstrated by their capacity to differentiate into adipocytic, osteocytic and chondrocytic lineages. Clonogenicity and cell proliferation ability were also tested. Furthermore, gene expression analysis by RT‐PCR was used to compare the expression of a set of genes (CD44, CD29, CD34, CD45, CD90, CD13, CD133, CD73, CD31 CD105, Oct4) with adipose tissue‐derived MSC. Stromal cells isolated from uterine endometrium showed similar morphology, ability of subculture and plasticity, and also expressed a panel of genes comparable with adipose tissue‐derived MSC. These data suggest that endometrial stromal cells fulfil the basic criteria proposed by the “Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy” for the identification of mesenchymal stem cells. Although endometrial mesenchymal stem cells (EnMSC) showed a lower replicative ability in comparison with adipose tissue‐derived MSC, they could be considered a cell therapeutic agent alternative to adipose tissue or bone marrow‐derived MSC in dog.     

Presence of opioid growth factor and its receptor in the normal dog,cat and horse cornea

Objective To determine if opioid growth factor (OGF, [Met⁵]enkephalin) and its specific receptor (OGFr) are present in normal cat, dog and horse cornea. Animals studied Normal dog, cat and horse. Procedure Corneas were obtained from animals euthanized for reasons unrelated to this project. One cornea from each of three normal cats, dogs and horses was evaluated. The right or left cornea from each animal was chosen randomly. Corneas were harvested and placed in corneal storage media for transport to The M.S. Hershey Medical Center of The Pennsylvania State University where immunocytochemistry techniques were used to demonstrate the presence and location of OGF and OGFr. Tissues were rinsed in Sorenson's phosphate buffer, immersed in 20% sucrose in buffer and then snap frozen in isopentane. Corneas were then embedded in OCT medium and 15 µm cryostat sections were created. Presence of OGF was determined by using a polyclonal antibody to [Met⁵]enkephalin and assessing immunoreactivity. OGFr presence was determined by using a previously characterized rabbit polyclonal antibody to the receptor. Results OGF and OGFr were identified in large quantities in the corneal epithelium of all three species. Conclusion Opioid growth factor and its specific receptor are present in the corneal epithelium of normal cats, dogs and horses. OGF is present in the cornea of many species and its presence is theorized to inhibit healing of injured tissue.     

Role of gap junctional intercellular communication (GJIC) through p38 and ERK1/2 pathway in the differentiation of rat neuronal stem cells

Gap junctional intercellular communications (GJIC) contributes to neural function in development and differentiation of CNS. In this study, we have investigated the expression of GJIC during the differentiation of neuronal stem cells and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced neuronal stem cell-derived cells from rat brain. During neuronal stem cell differentiation, expressions of Cx43 and 32 were increased for the duration of 72 hr, however the effect were decreased on the 7d. In the neuronal stem cell-derived cells, pretreatments with p38 MAP kinase inhibitor, SB203580, and MEK inhibitor, PD98059, could protect GJIC against TPA-induced inhibition of GJIC. Our data suggest that GJIC plays an important role during neuronal stem cell differentiation, and ERK1/2 and p38 MAP kinase signaling pathway may be closely related functionally to regulate gap junction in rat neuronal stem cell-derived cells.     

Corneal vascularization in the Florida manatee (Trichechus manatus latirostris) and three-dimensional reconstruction of vessels

The cornea of the Florida manatee is unique and unusual in its anatomy in that blood vessels have been found throughout. In all other animal species, this is considered a pathological condition impeding vision, and is usually caused by injury or trauma. The purpose of this study was to more clearly describe corneal vascularization by examining the architecture through three-dimensional reconstruction in order to find possible patterns in size, distribution, and location of blood vessels relative to gender, age, location, and season. Twenty-six eyes from 22 individuals were prepared for histologic examination and subsequent three-dimensional reconstruction. Every specimen examined had blood vessels in the cornea, comprising an average of 0.3% of total surface density (volume) of the cornea. No differences were found between individuals based on gender, age, and season. Environmental influences were not a significant factor either, which was not originally anticipated. The presence of vessels at the level of the anterior epithelium was surprising and it appeared that the vascularization was directed more anteriorly than was originally thought. The presence of blood vessels in a prenatal eye was also found. In all the eyes examined, no signs of injury or trauma could be observed. The presence of blood vessels appears to minimally impair vision based on their low density, size, and location. The association of vessels with the anterior epithelium and development of vessels within the fetus point to an evolutionary adaptation possibly due to the manatee's unique ability to move between water bodies.     

Bovine paramphistomosis in Galicia (Spain): Prevalence,intensity, aetiology and geospatial distribution of the infection

The present study explored various basic aspects of the epidemiology of paramphistomosis in Galicia, the main cattle producing region in Spain. In total, 589 cows from different farms located across the region were selected at random in the slaughterhouse for examination of the rumens and reticula for the presence of Paramphistomidae flukes. Paramphistomes were found in 111 of 589 necropsied cows (18.8%; 95% CI: 15.7–21.9%), with higher prevalences of infection in beef cows than in dairy cows (29.2% vs 13.9%). Although the number of flukes per animal was generally low (median = 266 flukes), some cows harboured large parasite burdens (up to 11,895 flukes), which may have harmful effects on their health or productivity. Cows with higher parasite burdens also excreted greater numbers of fluke eggs in their faeces, which suggests that heavily parasitized mature cows play an important role in the transmission of paramphistomosis. This role may be particularly important in Galicia, where the roe deer, which is the only wild ruminant in the study area, was found not to be a reservoir for the infection. The use of morpho-anatomical and molecular techniques applied to a large number of fluke specimens provided reliable confirmation that Calicophoron daubneyi is the only species of the family Paramphistomidae that parasitizes cattle in Galicia. The environmental data from the farms of origin of the necropsied cows were used in Bayesian geostatistical models to predict the probability of infection by C. daubneyi throughout the region. The results revealed the role of environmental risk factors in explaining the geographical heterogeneity in the probability of infection in beef and dairy cattle. These explanatory factors were used to construct predictive maps showing the areas with the highest predicted risk of infection as well as the uncertainty associated with the predictions.     

Amplification of feline sarcoid‐associated papillomavirus DNA sequences from bovine skin

Feline sarcoids are uncommon dermal neoplasms that are thought to be caused by papillomaviral (PV) infection. Feline sarcoid‐associated PV (FeSarPV) has been consistently detected in sarcoids from North American and New Zealand cats but has not been detected within any other feline sample. This suggests that feline sarcoids may develop due to cross‐species infection by a PV from an unidentified reservoir host. While there is some epidemiological evidence to suggest that cattle are the reservoir host of FeSarPV, this PV has never been identified within any bovine sample. In this study both consensus PCR primers and primers specific to FeSarPV were used to investigate the presence of PV DNA within five fibropapillomas and 18 samples of inflammatory skin disease from cattle. Consensus primers amplified bovine PV‐2 DNA from four fibropapillomas, but none of the dermatitis samples. However, specific primers amplified FeSarPV DNA from four fibropapillomas and five inflammatory skin lesions. To the best of our knowledge this is the first time that FeSarPV has been detected within any sample other than a feline sarcoid. The ability of FeSarPV to asymptomatically infect bovine skin suggests that cattle are the reservoir host of this PV and feline sarcoids could be the result of cross‐species infection of a dead‐end host by a bovine PV.     

Hormographiella aspergillata keratomycosis in a dog

A 4-year-old, female, Border Collie was presented to the University of Bern Veterinary Teaching Hospital, because of a corneal lesion of 10 days duration. The axial cornea presented a whitish fluorescein-positive plaque with irregular margins. A diagnosis of keratomycosis was made based on cytology. Medical therapy with local broad-spectrum antibiotic and fluconazole was instituted. After 1 week of treatment, the improvement was deemed unsatisfactory. Therefore, a lamellar keratectomy and conjunctival pedicle flap were performed. After surgery, the cornea healed uneventfully. Histology confirmed the diagnosis of keratomycosis. The fungus could not be grown in culture and a precise etiological diagnosis could only be obtained with genetic identification of the fungus. A PCR technique was used to amplify the fungal genome from the cornea. Hormographiella aspergillata , the asexual reproductive form of the basidiomycete Coprinopsis cinerea , was identified. As advised in human medicine, we encourage the use of this molecular technique to obtain an early species diagnosis, allowing targeted medical therapy.     

Mycoplasmal mastitis in dairy cows in the Moghan region of Ardabil State, Iran

Mycoplasmas are an important and economically significant cause of mastitis in dairy cows in various parts of the world. The organisms are highly contagious, with the main reservoir of infection originating from cows with subclinical mastitis. In 1998 the 1st cases of bovine mastitis due to Mycoplasma bovis were diagnosed in Ardabil State, Iran. An investigation was carried out with the aim of establishing the extent of mycoplasma infections in dairy cows in Ardabil State. Milk samples obtained from 80 cows with clinical mastitis were cultured in the laboratory for the presence of mycoplasmas. Similarly, 48 bulk-tank milk samples were examined for the presence of mycoplasmas. A modified Hayflick broth was used to isolate the mycoplasmas and an immunoperoxidase test used for the species identification of the isolates. Mycoplasma bovis was isolated from 39 (48.75%) of the clinical mastitis samples and from 48 of the bulk-tank milk samples tested. This indicated that mycoplasma udder infections were more prevalent in dairy cows in Ardabil State than previously thought.     

Rodent hosts and flea vectors in Brazilian plague foci: a review

Plague, caused by the Yersinia pestis bacterium, has several foci scattered throughout a large area from the Brazilian territory that ranges from the Northeastern State of Ceará to the Southeastern State of Minas Gerais and another separated area at the State of Rio de Janeiro. This review gathers data from plague control and surveillance programs on the occurrence and geographic distribution of rodent hosts and flea vectors in the Brazilian plague areas during the period of from 1952 to 2019. Furthermore, we discuss how the interaction between Y. pestis and some rodent host species may play a role in the disease dynamics. The absence of human cases nowadays in Brazil does not mean that it was eradicated. The dynamics of plague in Brazil and in other countries where it was introduced during the 3rd pandemic are quite alike, alternating epidemics with decades of quiescence. Hence, it remains an important epidemic disease of global concern. The existence of a large animal reservoir and competent vectors demonstrate a need for continuous surveillance to prevent new outbreaks of this disease in humans.     

The high prevalence of hepatitis E virus infection in wild boars in Ibaraki Prefecture,Japan

Hepatitis E virus (HEV) is known as a causative agent of zoonosis and food poisoning. Pigs and some species of wild animals, including wild boar, are known to be a reservoir of HEV. In this study, we investigated the situation regarding HEV infection in wild boars in Ibaraki Prefecture, Japan. Serum, liver and feces samples from 68 animals were collected, and the presence or absence of HEV genomic RNA and HEV antibodies were analyzed. The viral genome was detected in samples from 7 (10.3%) animals, with all HEVs classified as genotype 3, subtype 3b. HEV antibodies were detected in samples from 28 (41%) animals. This report demonstrates for the first time the high prevalence of HEV infection in wild boars in Ibaraki Prefecture.     

鸡胚胎干细胞及其应用研究进展

鸡胚胎干细胞是一种多能性干细胞,从X期胚盘分离胚盘细胞或早期鸡胚的生殖嵴分离原始生殖细胞,经体外长期抑制分化培养可得到鸡胚胎干细胞。为维持细胞在培养过程中的未分化状态,需要采用饲养层细胞培养,同时设计合理的培养液配方并添加多种抑制分化或促进增殖的细胞因子。通过碱性磷酸酶活性检测、胚胎表面特异性抗原检测、分化试验及嵌合体试验等方法,可对鸡胚胎干细胞进行准确鉴定。文章主要就鸡胚胎干细胞的分离、培养与鉴定方法的研究进展及其应用前景进行简要综述,为进一步发展更高效的鸡胚胎干细胞培养体系并应用于生产实践提供一定的借鉴。     

Occurrence of gastrointestinal parasites in horses in metropolitan Perth, Western Australia

OBJECTIVE: To assess the occurrence of gastrointestinal parasites in horses in Perth. To apply polymerase chain reaction (PCR) for the identification of some species of encysted larval cyathostomes. DESIGN: Between February and September of 2000, the gastrointestinal tracts of 29 horses submitted to a local knackery and Murdoch University Veterinary hospital in Perth were examined post mortem for the presence of gastrointestinal parasites. PROCEDURE: The gastrointestinal tract was divided into six sections, which were screened for the presence of parasites such as Gasterophilus sp, Anoplocephala sp and Parascaris equorum. Samples of contents were taken for worm counts. RESULTS: Cyathostomes were found in 28 of the 29 horses. Eighteen species of gastrointestinal helminths were identified. Twelve of these were cyathostomes, with the four most common species being Cyathostomum catinatum, Cylicocyclus nassatus, Cylicostephanus longibursatus and Cylicostephanus goldi. The large strongyle, Triodontophorus serratus, was found in three of the horses but species of Strongylus were not found. CONCLUSIONS: In contrast to a study conducted on horses from this region in 1985, cyathostomes were the most common gastrointestinal parasites found. The four most common cyathostome species found in the present study correlated well with findings of studies in other locations. The high number of cyathostomes may be due to the increase in resistance to anthelmintics among the species, and to improper anthelmintic use. The apparent reduction in number of large strongyles may be due to the widespread use of ivermectin, which is very effective against these parasites, and also possibly because some larvae may not have been detected.