Visualization of Surface-specific Antigens in Various Strains of Enterotoxigenic E. coli

Proteinaceous surface antigens of enterotoxigenic E. coli (ETEC) appear as pili, and are important virulence factors as they allow bacteria to attach to the small intestinal mucosa. Surface antigens are classified as colonization factor antigens (CFA) and coli surface antigens (CS). Known groups include CFA/I, CFA/II (consisting of CS1, CS2 and CS3), CA/III and CFA/IV (consisting of CS4, CS5 and CS6). The goal of the present study was to examine the morphology of pili by transmission electron microscopy (TEM) and to localize specific surface antigens by immunolabelling. Using different strains of E. coli grown under various culture conditions, pili were visualized by negative staining and corresponding surface antigens were demonstrated by immunogold-labelling using both polyclonal and monoclonal antibodies. Expression of pili was dependent on culture conditions and sample handling. In contrast to CFA/I and CS3, CS6 pili were not detectable after negative staining. Selected antibodies, however, allowed surface antigens to be demonstrated unequivocally. These results will be of value in investigating the expression of colonizing factors in genetically modified bacterial strains.     

微生态制剂调控比格幼犬生长性能和肠道菌群的研究

【目的】 研究复合微生态制剂对哺乳期比格幼犬生长性能与肠道菌群结构的影响。【方法】 选取12只15日龄比格犬, 随机分为对照组和微生态制剂组, 每组6只。微生态制剂组幼犬每日灌喂犬用微生态制剂20 mL, 对照组幼犬灌喂生理盐水20 mL, 于试验第0、3、9、12、15、18、21、25、28 d测量体重并计算净增重, 第28天采集全血样、血清样和粪样。全血样进行红细胞(RBC)计数和血红蛋白(HGB)浓度测定, 血清样检测碱性磷酸酶(AKP)、无机磷(IP)、总胆固醇(TCHO)、甘油三酯(TG)、免疫球蛋白A(IgA)、免疫球蛋白G(IgG)、白介素-1β(IL-1β)、脂多糖(LPS)含量; 粪样进行菌群测序, 统计Goods coverage指数、Observed species指数和Shannon指数, 对样本进行PCoA分析, 并统计各样本在门、属、种水平上的物种分布。【结果】 与对照组相比, 微生态制剂可显著提高幼犬体重(P<0.05), 显著增加血清中RBC数量及HGB、AKP和IP含量(P<0.05), 促进比格幼犬生长发育; 微生态制剂可显著降低血清TG含量(P<0.05), 但对TCHO、IgA、IgG、IL-1β、LPS含量无显著影响(P>0.05)。Observed species指数和PCoA分析结果表明, 微生态制剂可显著改变幼犬肠道菌群结构(P<0.05)。在门水平上, 微生态制剂组幼犬肠道菌群拟杆菌门和放线菌门相对丰度显著降低(P<0.05), 梭菌门、酸杆菌门和疣微菌门相对丰度均显著增高(P<0.05);在属水平上, Escherichia-Shigella、霍尔德曼氏菌属、双歧杆菌属和布劳特氏菌属相对丰度均显著降低(P<0.05), 梭菌属、链球菌属、梭杆菌属、乳球菌属和片球菌属相对丰度均显著增高(P<0.05)。在种水平上, 大肠杆菌和短双歧杆菌相对丰度均显著降低(P<0.05), 犬大肠梭菌、格氏乳球菌、Vagococcus teuberi、贝莱斯芽孢杆菌、粪肠球菌和死亡梭杆菌相对丰度均显著增高(P<0.05)。【结论】 犬用微生态制剂可显著促进幼犬生长, 提高其血氧含量, 改变肠道菌群结构, 优化肠道菌群组成。     

The embryonic development of the bovine stomach revisited

The adult anatomy and physiology of the bovine (Bos taurus) stomach have been investigated extensively. Despite the many studies, however, the early development of the stomach has not yet been fully elucidated. The goal of the present study, therefore, was to review the available literature, to visualize the embryonic and early foetal development of the bovine stomach and to shed light on unresolved issues. The stomachs of fifteen bovine embryos and eleven foetuses from 26 to 80 days of gestation were photographed both in situ and after exenteration and critical point drying. A series of photographs was obtained that yielded a contiguous and comprehensive view of all the developmental changes that occurred until the virtually final configuration of the stomach was attained. In addition, the serosal surface was studied by electron microscopy, thus revealing subtle regional differences in the lining of the peritoneal cavity. Our observations corroborate the contention that all the compartments evolve from the fusiform primordium and that no outgrowth at the level of the oesophagus occurs. The greater curvature as well as the attachment line of the dorsal mesogastrium shift to the left, which is similar to the process in monogastrians. The rumen and reticulum develop from separate protrusions, and further compartmentalization results from constrictions and bulges and not from folding. Between 55 and 60 days of gestation, the entire bovine stomach except for the abomasum eventually relocates to its final position. In summary, previously debated key issues were addressed and integrated with current findings.     

Intestinal atresia and ectopia in a bovine fetus

A 2-year-old Red Holstein cow was presented with uterine torsion at 235 days of pregnancy. The fetus extracted by cesarean section had weak vital signs and marked abdominal distention. An edematous pouch that contained tubular structures with peristaltic activity was associated with the umbilical cord. Because of poor prognosis, both dam and fetus were euthanized. At necropsy, the fetus had severe distention of the forestomachs, abomasum, and proximal small intestine; absence of distal small intestine, cecum, and proximal colon; atresia of the 2 blind ends of the intestine; and atrophy of distal colon and rectum. The tubular structures associated with the umbilical cord were identified as the segments of intestine that were absent in the fetus. Intestinal atresia combined with ectopia may be caused by local ischemia during temporary herniation and rotation of the fetal gut into the extraembryonic coelom. The close connection between ectopic intestine and amniotic sheath of the umbilical cord in this case may have facilitated vascularization and allowed development and viability of the ectopic intestine.     

Ultrasound‐guided proximal paravertebral anaesthesia in cattle

ObjectiveTo develop and evaluate a method for ultrasound-guidance in performing the proximal paravertebral block for flank anaesthesia in cattle through a cadaveric study, followed by clinical application.Study designprospective experimental cadaveric study and clinical series.AnimalsPreviously frozen lumbar sections of cows without known spinal abnormalities were used. The clinical case group comprised of ten animals for which a right flank laparotomy was indicated.MethodsTwenty cow cadavers were used to perform ultrasound-guided bilateral injections of 1.0 mL dye (1.0 mL 1% Toluidine Blue in 1% Borax) at the intervertebral foramen at the level of T13, L1 and L2 spinal nerves. Distance and depth of injection, staining of the dorsal and ventral nerve branches, and deviation from the target were evaluated. The investigator’s confidence as to visualisation and expected success at staining the nerve was assessed. Ten clinical cases received the ultrasound-guided proximal paravertebral anaesthesia. Analgesic success was evaluated using a 4-grade scoring system at 10 minutes after the injection and during surgery, respectively. Categorical variables were described using frequencies and proportions.ResultsBoth dorsal and ventral branches of the spinal nerves T13, L1 or L2 were at least partially stained in 41% of injections, while in 77% of injections one of the branches was stained. Five out of ten clinical cases had a satisfactory anaesthesia. There was no significant association between confidence at injection and either staining or analgesic success.ConclusionResults from the cadaveric and clinical study suggest no significant improvement using ultrasound guidance to perform proximal paravertebral block in cows compared to our previous clinical experience and to references in the literature using the blind method.Clinical relevanceFurther research should be conducted to improve the ultrasound-guided technique described in this study.     

Primary ciliary dyskinesia with situs inversus totalis, hydrocephalus internus and cardiac malformations in a dog

A nine-month-old golden retriever bitch was presented with exercise intolerance and recurrent nasal discharge. Based on clinical, radiographic and ultrasonographic examination, a diagnosis of rhinitis, situs inversus totalis and tricuspid valve insufficiency was established. The results of video- and electron microscopy studies of the respiratory epithelium were compatible with primary ciliary dyskinesia (PCD). However, no evidence of a primary ultrastructural defect of the cilia was found. The dog was euthanased because of the poor prognosis. At necropsy, a hydrocephalus internus and a subaortic stenosis were additionally diagnosed. PCD, in combination with situs inversus, has been previously reported in golden retrievers, but without a concomitant hydrocephalus internus. Furthermore, concomitant occurrence of internal cardiac malformation and PCD has not previously been reported in the dog.     

Immunhistochemical Demonstration of Lumbar Intervertebral Disc Innervation in the Dog

Low back pain is a common ailment in dogs, particularly in specific breeds as for instance the German shepherd dog. A number of structures such as facet joint capsules, ligaments, dorsal root ganglia, periosteum, vertebral endplate, and meninges have been associated with this condition. However, in a substantial proportion of all cases, the origin of pain remains obscure. A further structure often being involved in vertebral column disorders is the intervertebral disc. However, the presence of nerves is a precondition for pain sensation and, consequently, structures lacking sensory innervation can be left out of consideration as a cause for low back pain. Nerve fibres have been demonstrated at the periphery of intervertebral discs in man, rabbit and rat. With regard to the dog, however, extent of intervertebral disc innervation still is disputed. The goal of the present study, therefore, was to assess previous findings based on silver impregnation. Protein gene product (PGP) 9.5 was used as a marker for immunohistochemical examination of serial transversal and sagittal paraffin sections of lumbar discs from adult dogs. This technique revealed nerve fibres to be confined to the periphery of the intervertebral discs. Thus, even limited pathological processes affecting the outer layers of the intervertebral disc are prone to cause low back pain.     

Effects of stimulus with proinflammatory mediators on nitric oxide production and matrix metalloproteinase activity in explants of cranial cruciate ligaments obtained from dogs

OBJECTIVE: To evaluate the origin and degree of activity of nitric oxide (NO) and matrix metalloproteinase (MMP) in explants of cranial cruciate ligaments (CCLs) obtained from dogs and cultured with and without inflammatory activators. SAMPLE POPULATION: Tissue specimens obtained from 7 healthy adult Beagles that were (mean +/- SD) 4.5 +/- 0.5 years old and weighed 12.5 +/- 0.8 kg. PROCEDURE: The CCLs were harvested immediately after dogs were euthanatized, and specimens were submitted for explant culture. Cultures were stimulated by incubation with a combination of interleukin-1, tumor necrosis factor-alpha, and lipopolysaccharide, or they were not stimulated. Culture supernatants were examined for production of NO nitrite-nitrate metabolites (NOts) and activity of MMP Cultured specimens were evaluated by use of immunohistochemical analysis to detect activity of inducible NO synthase (iNOS). RESULTS: All ligament explants produced measurable amounts of NOts. Stimulated cultures produced significantly more NOts after incubation for 24 and 48 hours, compared with nonstimulated cultures. Production of MMP in supernatants after incubation for 48 hours was significantly higher in stimulated cultures than in nonstimulated cultures. Cells with positive staining for iNOS were detected on all slides. Positively stained cells were predominantly chondroid metaplastic. There was a significant difference in intensity of cell staining between stimulated and non-stimulated cultures. CONCLUSIONS AND CLINICAL RELEVANCE: Explant cultures of intact CCLs obtained from dogs produce iNOS-induced NO. Stimulation of chondroid metaplastic cells in CCL of dogs by use of inflammatory activators can increase production of iNOS, NOts, and MMP.