An oil-emulsion vaccine induces full-protection against Mycoplasma agalactiae infection in sheep

The immunogenicity and efficacy of three inactivated vaccines (A, B, C) prepared with Mycoplasma agalactiae (M. agalactiae) and with different oil-emulsion adjuvants were evaluated in sheep. Twenty-eight animals were used, divided into four groups (a, b, c, d) of seven animals each. Three groups were immunized with the same vaccine, but using different adjuvants, while one group (d) was used as an unvaccinated control group. All the vaccine formulations were able to induce clinical protection of animals after challenge with M. agalactiae, but only vaccine C, emulsioned with Montanide ISA-563, Marcol-52 and Montane-80 (ratio: 30%, 63%, 7% respectively), was able to induce full protection in challenged animals, preventing both the onset of clinical signs and infection by M. agalactiae.     

Inactivated vaccine induces protection against Mycoplasma agalactiae infection in sheep

The efficacy of an inactivated oil-emulsion vaccine against Mycoplasma agalactiae was evaluated by an experimental infection of sheep. The vaccinated sheep developed high levels of antibodies and, following challenge, they did not develop any clinical signs of disease and the mycoplasmas were not detected, either by isolation trials or PCR assays carried out both on nasal swabs and milk specimens. The unvaccinated-challenged sheep showed typical signs of M. agalactiae infection and bacterial shedding. The results obtained indicate a good efficacy of the vaccine in eliciting protection against M. agalactiae infection.     

Passive immunization of Nile tilapia (Oreochromis niloticus) provides significant protection against Streptococcus agalactiae

A study was conducted to determine the role of specific antibodies in immunity to Streptococcus agalactiae. Adult Nile tilapia (Oreochromis niloticus) were injected i.p. with tryptic soy broth as control or with S. agalactiae vaccine. Ninety days later, fish were challenged with 1.5x10(4)CFUS. agalactiae fish(-1). Blood was drawn from all fish 90d after vaccination and 25d after challenge, and the acquired serum was injected i.p. in fingerling Nile tilapia. These passively immunized fish were subsequently challenged 72h later with 1.5x10(4)CFUS. agalactiae fish(-1), and significantly less (P<0.0001) mortalities were noted among fish administered serum containing specific anti-S. agalactiae antibodies (0.0-10.0% mortalities) than in control groups (63.3-72.7% mortalities). Heat-inactivation of serum produced no significant differences in mortalities than non-heat-treated serum in groups administered serum containing specific antibodies from vaccinated fish (P<0.9455) or vaccinated-challenged fish (P<0.0781). Pre-challenge serum samples indicate that the passively immunized fish had significantly increased (P<0.0001) specific antibody levels over control fish. A highly significant (r(2)=0.5892; P<0.0001) correlation between increased pre-challenge specific serum antibody OD levels and survival after challenge was demonstrated when analyzing the control and passive immunization groups. The results of this study indicate that specific anti-S. agalactiae antibodies play a primary role in immunity to S. agalactiae in fish.     

A chromosomal region of Mycoplasma agalactiae containing vsp-related genes undergoes in vivo rearrangement in naturally infected animals

A Mycoplasma agalactiae genomic fragment carrying four vsp-related genes (designated avg: agalactiae variable genes) was cloned, sequenced and compared to the vspA gene of Mycoplasma bovis. The following features were revealed: (i) the presence of a highly conserved vsp 5' upstream region; (ii) a highly homologous vsp N-terminal end encoding a putative lipoprotein signal sequence; (iii) sequence divergence of the rest of the mature proteins. By using avg specific probes in Southern blot analysis of genomic DNAs of M. agalactiae strains as well as of isolates from infected animals, marked DNA polymorphism of avg fragments was demonstrated. In addition, the avg genomic fingerprints were monitored for a period of 7 months, in isolates of M. agalactiae from an individual chronically infected animal. The results provided evidence that a chromosomal region of M. agalactiae, carrying vsp-related genes, undergoes rearrangements in vivo in the natural animal host during the course of infection.     

Potential pathogens carried by Spanish ibex (Capra pyrenaica hispanica) in southern Spain

The Spanish ibex (Capra pyrenaica hispanica) population of southern Spain was surveyed for potential pathogens associated with the conjunctiva, external ear canal, as well as reproductive and upper respiratory tracts. We sampled 321 ibex (131 adult males, 100 adult females, and 90 yearlings); these included 271 apparently healthy animals and 50 that were naturally infected with Sarcoptes scabiei. A total of 688 bacterial isolates were identified (377 gram-negatives, 225 gram-positives, and 86 Mycoplasma spp.); sex, age, location, infection with S. scabiei, and disposition of the animal (free-ranging versus captive) were evaluated as risk factors for infection. Infections with Mycoplasma agalactiae and Mycoplasma arginini were associated with age, having a higher frequency of isolation in young animals. With Escherichia coli, Mannheimia haemolytica, Pasteurella multocida biotype A, and Staphylococcus aureus, significantly higher isolation rates were associated with adults. The isolation frequency for E. coli was higher in females, whereas Moraxella bovis isolations were mostly associated with males. The presence of mange increased the risk of infection with both Streptococcus equi subsp. zooepidemicus and M. haemolytica. The geographic origin of sampled animals was related to the isolation of Branhamella ovis, M. agalactiae, and all Pasteurella sp. Isolations of M. haemolytica, P. multocida biotype A, E. coli, and B. ovis were more prevalent in samples from free-ranging rather than captive animals. Of the gram-positive bacteria, S. aureus represented the predominant species isolated from nasal, vaginal, and ocular samples. Mycoplasma agalactiae and M. arginini were the predominant Mycoplasma spp., and both were associated most often with the external ear canal. The most frequently isolated gram-negative bacteria included E. coli, M. haemolytica, P. multocida biotype A, and B. ovis. Isolation rates of gram-negative species varied by source. In nasal samples, M. haemolytica and P. multocida biotype A were isolated most frequently, whereas in ocular and vaginal samples, B. ovis and E. coli, respectively, were most frequently isolated.     

Re-exposure of captive house finches that recovered from Mycoplasma gallisepticum infection

Fourteen house finches were reinoculated (re-exposed) with 0.05 ml (3.24x10(5) colony forming units/ml) of Mycoplasma gallisepticum (MG) in the conjunctival sac of each eye. All birds used in this reinoculation study had recovered from previous infection between 27 and 83 days after inoculation. Recovery was based on the absence of clinical signs of conjunctivitis and/ or the inability to detect MG in conjunctival or choanal samples. Birds were maintained in individual cages under controlled environmental conditions at temperature 21-24 C, relative humidity 70%, and a light cycle adjusted to ambient values. They were divided into three groups, (A, B, and C). Five birds each were reinoculated 219 days (7.3 mo, group A) and 314 days (10.47 mo, group B) after the original infection. The final group of four birds was reinoculated at 425 days after experimental infection (14.17 mo, group C). Although the birds were randomly assigned to the three groups, the duration of the disease state (number of days until clinical signs last observed) during initial infection differed: group A mean=37.0+/-SE 4.549, group B mean=63.6+/-SE 6.306, group C mean=42.75+/-SE 2.750; analysis of variance F2,11=8.17, P=0.007. Within 24 hr after reinoculation six of the 14 experimental birds had developed some clinical signs of MG-induced conjunctivitis. At 3 days after reinoculation, 12 of the 14 birds had unilateral or bilateral conjunctivitis. The duration of clinical signs in the reinoculated individuals was significantly shorter than with their previous infection. These results suggest that the birds were able to mount a rapid and strong immune response following re-exposure. However, they were susceptible to reinfection and developed disease, suggesting that reinfection or perhaps even recurrence of infection and disease could occur in the free-ranging population. This may represent an important component in the epidemiology of this disease in house finches.     

Further studies on an intermediate host murine model showing that a primary Echinococcus granulosus infection is protective against subsequent oncospheral challenge

We describe the use of a murine model to evaluate resistance against subsequent challenge following a primary infection with oncospheres of Echinococcus granulosus. Mice (Kunming strain) were infected with hatched oncospheres of Echinococcus granulosus; 21 days later a second challenge was given by a different route of infection. A primary infection by intraperitoneal (i.p.) injection stimulated 100 and 90.5% protection in terms of reduced cyst numbers against a secondary infection given subcutaneously (s.c.) or intravenously (i.v.), respectively. A primary infection given s.c. followed by i.p. or i.v. challenge resulted in 84.0 and 100% protection, respectively. Intravenous infection followed by i.p. or s.c. challenge resulted in 98.5 and 69.4% protection, respectively. With the i.v. route of infection, almost all resultant cysts were present in the lungs. The data show that a primary infection with oncospheres can induce total or a high degree of protection against a subsequent challenge and confirms that natural (concomitant) immunity can be stimulated in the intermediate host as the result of a primary infection. This may explain the decline in hydatid infection in sheep older than 2 years in hyper-endemic areas such as those found in Xingjiang, China. These older sheep may have been earlier infected and have subsequently self-cured, with the primary infection stimulating an immune response that protects the intermediate host animals from further infection.     

Construction of the first shuttle vectors for gene cloning and homologous recombination in Mycoplasma agalactiae

Mycoplasma agalactiae is a worldwide ruminant pathogen that causes significant economic losses by inflicting contagious agalactia in sheep and goats. The development of efficient control strategies requires a better understanding of the mycoplasma factors that promote successful infection. However, lack of genetic tools has been a major impediment in studying the pathogenic mechanisms of M. agalactiae. This study describes the identification and cloning of the M. agalactiae origin of replication (oriC) in order to construct the first shuttle vectors for targeted gene disruption, gene complementation and expression studies. Additionally, this report provides the first evidence of the occurrence of homologous recombination and the functionality of heterologous tetM determinant in this pathogen.     

A real-time PCR assay for detection and quantification of Mycoplasma agalactiae DNA

AIMS: The aim of this study was to develop a rapid, sensitive, specific tool for detection and quantification of Mycoplasma agalactiae DNA in sheep milk samples. METHODS AND RESULTS: A real-time polymerase chain reaction (PCR) assay targeting the membrane-protein 81 gene of M. agalactiae was developed. The assay specifically detected M. agalactiae DNA without cross-amplification of other mycoplasmas and common pathogens of small ruminants. The method was reproducible and highly sensitive, providing precise quantification of M. agalactiae DNA over a range of nine orders of magnitude. Compared with an established PCR assay, the real-time PCR was one-log more sensitive, detecting as few as 10(1) DNA copies per 10 microl of plasmid template and 6.5x10(0) colour changing units of reference strain Ba/2. CONCLUSIONS: The real-time PCR assay is a reliable method for the detection and quantification of M. agalactiae DNA in sheep milk samples. The assay is more sensitive than gel-based PCR protocols and provides quantification of the M. agalactiae DNA contained in milk samples. The assay is also quicker than traditional culture methods (2-3 h compared with at least 1 week). SIGNIFICANCE AND IMPACT OF THE STUDY: The established real-time PCR assay will help study the patterns of shedding of M. agalactiae in milk, aiding pathogenesis and vaccine efficacy studies.     

Epidemiology of Mycoplasma agalactiae infection in free-ranging Spanish ibex (Capra pyrenaica) in Andalusia, southern Spain

Mycoplasma agalactiae is the main causal agent of contagious agalactia syndrome in Spain. It is a severe disease of small ruminants, endemic in Mediterranean countries, that is characterized by mastitis, arthritis, and keratoconjunctivitis. This paper investigates the temporal, spatial, and host-related factors in the distribution of M. agalactiae infection from October 1996 to November 1998 and March 2002 to May 2003 in Spanish ibex (Capra pyrenaica) populations from Andalusia, in southern Spain. The predisposing factors to infection among previously selected factors (year of sampling, climatic season, geographic origin according to province, mountain range and metapopulation, sex, year of life, presence of scabies, and phase of the reproductive cycle) were established. We collected conjunctival and ear-canal swabs from 411 free-ranging ibexes. The frequency of infected ibexes was 11.2%. The peak frequency of infection occurred in 1998 and in summer. Granada was the province with greatest risk (odds ratio = 2.6) of carriers (18.8% infected). The predisposing factors were sex (females), age (young animals), and metapopulation (Sierra Nevada). We identified a higher number of infected ibexes in the metapopulation "Sierra Nevada" (34/ 256) and significant differences among the three established metapopulations (P<0.01). Mycoplasma agalactiae infection represents a risk for population density and maintenance of these wild populations; infections can result in blindness, malnutrition, and polyarthritis leading to numerous deaths.     

Mycobacterium Avium subsp. Paratuberculosis Isolates Induce In Vitro Granuloma Formation and Show Successful Survival Phenotype,Common Anti-Inflammatory and Antiapoptotic Responses within Ovine Macrophages Regardless of Genotype or Host of Origin

The analysis of the early macrophage responses, including bacterial growth within macrophages, represents a powerful tool to characterize the virulence of clinical isolates of Mycobcaterium avium susbp. paratuberculosis (Map). The present study represents the first assessment of the intracellular behaviour in ovine monocyte-derived macrophages (MDMs) of Map isolates representing distinct genotypes (C, S and B), and isolated from cattle, sheep, goat, fallow deer, deer, and wild boar. Intracellular growth and survival of the selected isolates in ovine MDMs was assessed by quantification of CFUs inside of the host cells at 2 h p.i. (day 0) and 7 d p. i. using an automatic liquid culture system (Bactec MGIT 960). Variations in bacterial counts over 7 days from the baseline were small, in a range between 1.63 to 1.05-fold. After 7 d of infection, variations in the estimated log₁₀ CFUs between all the tested isolates were not statistically significant. In addition, ovine MDMs exhibited enhanced anti-inflammatory, antiapoptotic and antidestructive responses when infected with two ovine isolates of distinct genotype (C and S) or with two C-type isolates from distinct hosts (cattle and sheep); which correlated with the successful survival of these isolates within ovine MDMs. A second objective was to study, based on an in vitro granuloma model, latter stages of the infection by investigating the capacity of two Map isolates from cattle and sheep to trigger formation of microgranulomas. Upon 10 d p.i., both Map isolates were able to induce the formation of granulomas comparable to the granulomas observed in clinical specimens with respect to the cellular components involved. In summary, our results demonstrated that Map isolates from cattle, sheep, goats, deer, fallow-deer and wild boar were able not only to initiate but also to establish a successful infection in ovine macrophages regardless of genotype.     

Mycoplasma bovis shares insertion sequences with Mycoplasma agalactiae and Mycoplasma mycoides subsp. mycoides SC: Evolutionary and developmental aspects

Three new insertion elements, ISMbov1, ISMbov2 and ISMbov3, which are closely related to ISMag1 (Mycoplasma agalactiae), ISMmy1 and IS1634 (both Mycoplasma mycoides subsp. mycoides SC), respectively, have been discovered in Mycoplasma bovis, an important pathogen of cattle. Southern blotting showed that the genome of M. bovis harbours 6-12 copies of ISMbov1, 11-15 copies of ISMbov2 and 4-10 copies of ISMbov3, depending on the strain. A fourth insertion element, the IS30-like element, is present in 4-8 copies. This high number of IS elements in M. bovis, which represent a substantial part of its genome, and their relatedness with IS elements of both M. agalactiae and M. mycoides subsp. mycoides SC suggest the occurrence of two evolutionary events: (i) a divergent evolution into M. agalactiae and M. bovis upon infection of different hosts; (ii) a horizontal transfer of IS elements during co-infection with M. mycoides subsp. mycoides SC and M. bovis of a same bovine host.     

岩藻糖基化人乳低聚糖在新生儿无乳链球菌肺炎治疗中的作用

目的探讨岩藻糖基化人乳低聚糖在新生儿无乳链球菌肺炎中的治疗作用。方法收集本院2015年7月至2017年2月痰培养阳性新生儿无乳链球菌、大肠埃希菌和肺炎克雷伯菌感染性肺炎病例,分析不同喂养方式、不同病原类别肺炎在住院时间、炎症、并发症等指标差异,探讨岩藻糖基化人乳低聚糖对无乳链球菌感染的独特治疗作用;使用含有不同浓度岩藻糖基化人乳低聚糖的培养基对无乳链球菌进行培养,研究岩藻糖基化人乳低聚糖抑制无乳链球菌的最佳浓度;分别使用岩藻糖基化人乳低聚糖或酸性人乳低聚糖加入培养基中对无乳链球菌、大肠埃希菌、肺炎克雷伯菌进行培养,进一步探讨岩藻糖基化人乳低聚糖对无乳链球菌的独特抑制作用。结果无乳链球菌肺炎患儿,母乳喂养组的住院时间较人工喂养组住院时间减少[(8.35±0.77)d vs(10.91±0.54)d,t=1.557 676,P<0.05)],PCT值较人工喂养组低[(2.38±0.63)ng/mL vs(8.69±2.23)ng/mL,t=1.419 964,P<0.05],白细胞计数较人工喂养组低[(13.28±1.08)×10^9/L vs(16.16±0.98)×10^9/L,t=1.878 447,P<0.05],脑膜炎发生率较人工喂养组低(5%vs 35%,χ^2=5.601 353,P<0.05),呼吸机使用率较人工喂养组低(10.0%vs 36.4%,χ^2=4.005 042,P<0.05);大肠埃希菌肺炎患儿、肺炎克雷伯菌肺炎患儿,母乳喂养组与人工喂养组比较,住院时间稍减少,PCT值、白细胞计数、脑膜炎发生率和呼吸机使用率均有所降低,差异无统计学意义;细菌培养实验发现岩藻糖基化人乳低聚糖在2.5 mg/L浓度的时候可以明显抑制无乳链球菌的生长,对大肠埃希菌和肺炎克雷伯菌无明显抑制作用,酸性人乳低聚糖对3种细菌均无抑制作用。结论岩藻糖基化人乳低聚糖可以明显抑制无乳链球菌的生长,可能成为未来无乳链球菌感染性肺炎治疗中新的靶点,值得深入研究。     

Involvement of monoamine oxidase inhibition in neuroprotective and neurorestorative effects of Ginkgo biloba extract against MPTP-induced nigrostriatal dopaminergic toxicity in C57 mice.

The present study investigated the neuroprotective and neurorestorative effects of Ginkgo biloba extract (EGb 761) and its two components ginkgolides A (BN52020) and B (BN52021) in mice. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) (30 mg/kg/d i.p. for six days) significantly reduced striatal dopamine (DA) levels in C57 mice measured by high performance liquid chromatography with electrochemical detection (HPLC-EC). When C57 mice were pretreated with EGb 761 (20, 50, 100 mg/kg/d i.p.) for 7 days and then treated with the same extract 30 min before MPTP injection for 6 days, the neurotoxic effect of MPTP was antagonized in a dose-dependent fashion. Similar treatment with ginkgolides A and B (5, 10, 50 mg/kg/d i.p.) showed no protective effect. When C57 mice were treated with EGb 761 (50 mg/kg/d i.p.) after MPTP-lesion, the recovery of striatal dopamine (DA) levels was accelerated. However, similar treatment with ginkgolides A or B (10 mg/kg/d i.p.) did not show any effect. EGb 761, but not ginkgolides A and B, nonselectively inhibited mouse brain MAO activity in vitro (IC50 = 36.45 +/- 1.56 microg/ml) tested by an improved fluorimetric assay. The results demonstrate that EGb 761 administered before or after MPTP treatment effectively protects against MPTP-induced nigrostriatal dopaminergic neurotoxicity and that the inhibitory effect of EGb 761 on brain MAO may be involved in its neuroprotective effect.     

Effects of a 100 metacercarial cyst inoculum on the host-parasite relationship of Echinostoma caproni and ICR mice

The host-parasite relationship of a 100 metacercarial cyst inoculum of Echinostoma caproni in the ICR mouse was examined. Three groups of mice, A, B and C, each with six mice per group were used and all mice were necropsied at 14 days postinfection (p.i.), at which time the worms were ovigerous. Group A consisted of uninfected controls, whereas group B received 25 cysts per mouse (low dose) and group C received 100 cysts per mouse (high dose). There was no significant difference in food consumption between any of the groups from 0 to 14 days p.i. Control mice increased their body weight by 12%, group B by 5%, and group C showed a less than 1% increase in body weight between 0 and 14 days p.i. Echinostome parasitism caused a significant increase in the diameter of the mouse gut, with the gut of group C being more significantly dilated than that of either group A or B. The average worm recovery from group B was 20 worms per host, compared to 72 worms per host from group C. The mean wet and dry weights per worm from group B were 2.4 and 0.4 mg, respectively as compared to 0.6 and 0.2 mg respectively for group C. The mean number of uterine eggs per worm from group B was 180 compared to 125 for worms from group C. Worms from group C were more widely distributed in the small intestine than those from group B. Crowding effects associated with the high dose infection were clearly demonstrated in E. caproni from ICR mice.     

Survival of bighorn sheep (Ovis canadensis) commingled with domestic sheep (Ovis aries) in the absence of Mycoplasma ovipneumoniae

To test the hypothesis that Mycoplasma ovipneumoniae is an important agent of the bighorn sheep (Ovis canadensis) pneumonia that has previously inevitably followed experimental commingling with domestic sheep (Ovis aries), we commingled M. ovipneumoniae-free domestic and bighorn sheep (n=4 each). One bighorn sheep died with acute pneumonia 90 days after commingling, but the other three remained healthy for >100 days. This unprecedented survival rate is significantly different (P=0.002) from that of previous bighorn-domestic sheep contact studies but similar to (P>0.05) bighorn sheep survival following commingling with other ungulates. The absence of epizootic respiratory disease in this experiment supports the hypothesized role of M. ovipneumoniae as a key pathogen of epizootic pneumonia in bighorn sheep commingled with domestic sheep.     

夏氏疟原虫感染过程中调节性B细胞的表达变化

调节性B细胞(regulatory B cells,Bregs)是近年来发现的通过分泌IL-10发挥免疫调节作用的B细胞,其在疟疾免疫应答中的作用尚不清楚。利用血液阶段夏氏疟原虫(Plasmodium chabaudi AS,P.c AS)感染的BALB/c小鼠,观察了感染过程中Bregs数量变化及其表面分子表达情况。结果显示,BALB/c小鼠感染P.c AS后红细胞感染率逐渐升高,于感染后第9天达到30%,多数小鼠死亡。脾组织细胞因子IL-10 mRNA水平在感染后显著升高,感染后第5天达到高峰,感染后第8天仍为正常小鼠的10倍左右。CD4+细胞中IL-10+细胞百分比和绝对数量在感染后第5天达到峰值,于感染后第8天回落至正常水平;而CD19+细胞中IL-10+细胞(Bregs)百分比在感染后持续上升,在感染后第8天达到CD19+细胞的5%左右;感染后第5天,脾中CD4+IL-10+细胞绝对数量高于CD19+IL-10+细胞,至感染后第8天,CD19+IL-10+细胞绝对数量显著高于CD4+IL-10+细胞(P﹤0.01),约90%Bregs为CD5-细胞。结果提示,P.c AS感染过程中Bregs显著活化,是感染1周后IL-10的主要产生细胞。     

Effects of intrauterine infusion of Escherichia coli endotoxin in normal cows and in cows with endometritis induced by experimental infection with Streptococcus agalactiae

Two experiments were carried out to describe the effects of intrauterine infusion of Escherichia coli endotoxin on some aspects of nonspecific uterine defense mechanisms in healthy cyclic cows (Experiment 1) and in cows with induced endometritis by experimental infection with Streptococcus agalactiae (Experiment 2). In Experiments 1 and 2, the mean log(e) total white cell counts (>95% neutrophils) in the uterine flushing fluid of the endotoxin-treated group were significantly increased (P<0.01 and P<0.05, respectively). Streptococcus agalactiae was detected by the Latex Agglutination Test (LAT) in 47% of the samples from uteri experimentally infected with this organism; 12.5% were positive on culture, and only 10% were positive on both tests. With one exception, all the samples with a positive culture were positive to the LAT, but not all samples submitted to the LAT had positive culture. There was a significant (P<0.05) association between endotoxin treatment and the presence of infection detected by the LAT but not with that detected by culture (P>0.05) at 3 to 12 days post infection. Similar results were found at 8 to 16 days post infection. The infection disappeared from the endotoxin-treated group but not from the non-treated group 12 days after the induction of infection. It is concluded that intrauterine E. coli endotoxin infusion might provide an alternative treatment for those cows with endometritis that is refractory to conventional antimicrobial and hormonal therapy. It is also concluded that the LAT is an easier, quicker and more reliable method than bacterial culture for the detection of endometritis caused by Strep. agalactiae , and, possibly, such immunodiagnostic tests may be useful for the detection of other uterine infections.     

Source of cell injected is a critical factors for short and long engraftment in xeno-transplantation

Abstract. This study aims to investigate engraftment of human cord blood and foetal bone marrow stem cells after in utero transplantation via the intracoelomic route in the sheep. Here, we performed transplantation in 14 single and 1 twin sheep foetuses at 40–47 days of development, using a novel schedule for injection. (i) Single injection of CD34⁺ human cord blood stem cells via the coelomic route (from 10 to 50 × 10⁴) in seven single foetuses. (ii) Single injection of CD34⁺ foetal bone marrow stem cells via the intracoelomic route with further numbers of cells (20 × 10⁵ and 8 × 10⁵, respectively) in three single and in one twin foetuses. (iii) Double fractioned injection (20–30 × 10⁶) via the coelomic route and 20 × 10⁶ postnatally, intravenously, shortly after birth of CD3-depleted cord blood stem cells in four single foetuses. In the first group, three single foetuses showed human/sheep chimaerism at 1, 8 and 14 months after birth. In the second group, the twin foetuses showed human/sheep chimaerism at 1 month after birth. In the third group, only two out of four single foetuses that underwent transplantation showed chimaerism at 1 month. While foetal bone marrow stem cells showed good short-term engraftment (1 month after birth), cord blood stem cells were able to persist longer in the ovine recipients (at 1, 8 and 14 months after birth).     

肺炎支原体感染小鼠体内IL-6、IFN-γ、IL-5的水平及克拉霉素的影响

目的：研究肺炎支原体感染小鼠血清及支气管肺泡灌洗液中细胞因子的水平,了解克拉霉素治疗对细胞因子的影响。方法：将昆明小鼠分为感染组、正常组及药物治疗组,建立小鼠肺炎支原体感染模型成功后,药物组用克拉霉素（6g·kg^-1·d^-1）进行治疗,连续5天。在肺炎支原体感染模型建立后的第8天,检测三组小鼠的血清及支气管肺泡灌洗液中IL-6、IFN-γ、IL-5水平。结果-相比较于正常组,感染组小鼠支气管灌洗液和血清中的IL-6、IFN-γ、IL-5均显著升高（P〈0．05）。而克拉霉素的使用能使IL-6、IFN-γ降低（P〈0．05）,但对IL-5水平无影响。结论：肺炎支原体感染使小鼠IL-6、IFN-γ、IL-5水平增高,克拉霉素治疗有一定疗效。