Isolation of a plasmid from "canine" Staphylococcus epidermidis mediating constitutive resistance to macrolides and lincosamides

A small plasmid of 2.5 kB mediating constitutive resistance to macrolide-lincosamide-(ML)antibiotics could be detected in a “canine” Staphylococcus epidermidis-culture. This plasmid, designated as pSES 1, was identified by interspecies protoplast transformation into Staphylococcus aureus RN 4220. A detailed restriction map of pSES 1 could be constructed using the restriction endonucleases Acc I, Bcl I, Cfo I, Cla I, Hind III, Hinf I, Mbo I, Sst I and Taq I. This map allowed structural comparisons of pSES 1 with plasmids from “human” Staphylococcus- and Bacillus-species, also mediating macrolide-lincosamide resistance (ML^R). On the basis of its restriction map, pSES 1 proved to be similar to the plasmids pNE 131 from “human” S. epidermidis, pE 194 from “human” S. aureus and pIM 13 from B. subtilis.     

Application of the California mastitis test in intramammary Streptococcus agalactiae and Staphylococcus aureus infections of camels (Camelus dromedarius) in Kenya

A study was conducted on 207 lactating camels in six herds in Kenya to evaluate the California mastitis test (CMT) for the detection of intramammary infections (IMIs) caused by Streptococcus agalactiae and Staphylococcus aureus and to investigate the prevalence of both the pathogens in the camel udder. IMI with S. agalactiae was found in 12% of all camels sampled. IMI with S. aureus was present in 11% of all camels sampled. The herd-level prevalence of IMI varied between 0 and 50% for S. agalactiae and between 0 and 13% for S. aureus. Longitudinal observations over 10–12 months confirmed persistent infections for both pathogens. Observations in one herd suggested that camel pox was a contributing factor in spreading and exacerbating S. agalactiae udder infections.

The CMT had quarter-level sensitivities of 77 and 68% for S. agalactiae and S. aureus in camels, respectively. The CMT specificities were 91% for both the pathogens.     

Correlation between carcass conformation and fat cover degree, and muscle fatty acid profile of yearling bulls depending on breed and mh-genotype

The objective was to study the relationships between the actual European beef carcass classification scale, which classifies carcasses with regard to conformation and degree of fat cover scores, and muscle fat quality, depending on breed and mh-genotype. For this purpose samples from 100 yearling bulls from “Asturiana de los Valles” (24 AV(mh/mh), 26 AV(mh/+), 25 AV (+/+)) and “Asturiana de la Montaña” (25 AM) were analysed. The results of the study showed that breed or genotype affect carcass measurement scores and muscle fatty acid profile through its important effect on animal overall fatness. Homozygous double-muscled animals produced carcasses with high conformation and low intramuscular (IM) fat content. While early-maturing and rustic AM animals produced low carcass yield and high IM fat content. The other genotypes (mh/+, +/+) showed, in general, intermediate characteristics. Referring to correlations, carcass conformation was negatively related to saturated (SFA) (r = − 0.69, P < 0.001) and monounsaturated fatty acid (MUFA) (r = − 0.69, P < 0.001) groups, and positively to polyunsaturated (PUFA) (r = 0.72), n-6 (r = 0.72), n-3 (r = 0.71) and unsaturated fatty acid (UFA) (r = 0.69) groups, being all of them significant (P < 0.001). However, carcass degree of fat cover was positively related to SFA (r = 0.53, P < 0.001) and MUFA (r = 0.62, P < 0.001), and negatively to PUFA (r = − 0.61), n-6 (r = − 0.60), n-3 (r = − 0.62) and UFA (r = − 0.53) groups, being all of them significant. Moreover, simple and low-cost prediction equations were calculated for a rapid and sufficiently accurate fatty acid group (SFA, MUFA, PUFA, n-6, n-3, UFA) estimation (R² > 0.46, P < 0.001). In general, meat obtained from double-muscled animals display a more appropriate IM fatty acid profile from the nutritional point of view according to actual recommendations, but it could happen the disability of these lean animals to deposit sufficient IM fat to ensure consumer overall liking or acceptability.     

Microbial activity in the gut of piglets: I. Effect of prebiotic and probiotic supplementation

Four groups of six 21 days-old piglets were used to evaluate the effect of a prebiotic or probiotic on the intestinal fermentative activity. In each group, piglets received one of the following diets: basal diet (C); basal diet supplemented with xylo-oligosaccharide (C-XOS); basal diet supplemented with Saccharomyces cerevisiae (C-SC); and basal diet supplemented with xylo-oligosaccharide and S. cerevisiae (C-XOS + SC).

The short chain fatty acids in the colon of piglets were decreased with the inclusion of S. cerevisiae in the diet (P < 0.01). The xylanolytic activity was higher (P < 0.05) in the small intestine of piglets fed C-XOS + SC diet, but no significant differences were found in the caecum and colon. In the caecum contents, the cellulolytic activity was increased (P < 0.05) by the C-XOS and C-SC diets, but remained similar when the diet was supplemented with the two additives combined.     

Effect of feeding different levels of foliage of Moringa oleifera to creole dairy cows on intake, digestibility, milk production and composition

An experiment was conducted in Nicaragua to determine the effect of feeding different levels of foliage from Moringa oleifera Lam (synonym: Moringa pterygosperma Gaertner) to dairy cows on intake, digestibility, milk production and milk composition. The treatments were: Brachiaria brizantha hay ad libitum, either unsupplemented or supplemented with 2 kg or 3 kg of Moringa on a dry matter (DM) basis. Six Bos indicus cows of the Creole Reyna breed, with a mean body weight of 394 ± 24 kg were used in a replicated 3 × 3 Latin square design. Supplementation with Moringa increased (P < 0.05) DM intake from 8.5 to 10.2 and 11.0 kg DM day^− 1 and milk production from 3.1 to 4.9 and 5.1 kg day^− 1 for B. brizantha hay only and supplementation with 2 kg and 3 kg DM of Moringa, respectively. Milk fat, total solids and crude protein and organoleptic characteristics, smell, taste and colour, were not significantly different between the diets. Apparent digestibility coefficients of DM, OM, CP, NDF and ADF increased (P < 0.05) in the diets supplemented with Moringa compared with B. brizantha hay alone. The results showed that the inclusion of Moringa as a protein supplement to low quality diets improved DM intake and digestibility of the diet and increased milk production but did not affect milk composition.     

Phenotypic and genotypic traits of Staphylococcus aureus strains isolated from pig carcasses

A total of 142 S. aureus strains isolated from pig carcasses from abattoirs A (n = 98) and B (n = 44) were characterized by phenotypic and genotypic traits. Phenotypically, 96% showed yellow-pigmented colonies, 63% β/δ hemolysis, 85% were egg yolk-positive and 99% were positive for clumping factor/protein A. Only 25% of the strains were resistant to the antimicrobials tested (abattoir A: 19%; abattoir B: 39%), especially to penicillin and ampicillin. None of the strains harbored the mecA gene, which is conserved in methicillin-resistant S. aureus. The biofilm associated genes icaA, icaD and bap were present in 100%, 100% and 0% of the strains. Genes for staphylococcal enterotoxin (SE) were detected in 51% (abattoir A) and 14% of the strains (abattoir B). Among strains harboring SE genes (n = 56), 63%, 31%, 4% and 2% tested positive for seg/sei, seg, sei and sec, respectively. The amplification of the 3′ end of the coagulase gene (coa) yielded amplicons of 400, 436, 602, 682 or 776 bp. Coa restriction profile (CRP) analysis using HaeIII resulted in seven patterns (a–d, e1–e3). CRP (c) was detected most frequently at both abattoirs, whereas CRP (a) was restricted to abattoir A and CRP (e3) to abattoir B. In the slaughter process (abattoir B), (i) two CRPs (b and d) were only found before dehairing/singeing, and (ii) four CRPs (c, e1–e3) were identified throughout the process. The genotyping revealed a remarkable homogeneity in S. aureus strains from the two different abattoirs and the slaughter process stages. These results may be explained by the distribution of a limited number of S. aureus genotypes in the pig population. Moreover, as the predominant CRPs (c, e1–e3) persisted throughout the slaughter process in abattoir B, it may be hypothesized that these types are characterized by colonization advantages.     

Interpretation of California mastitis test scores using Staphylococcus aureus culture results for screening of subclinical mastitis in low yielding smallholder dairy cows in the Dar es Salaam region of Tanzania

Screening of subclinical mastitis under field conditions is done using the California mastitis test (CMT). CMT score of ≥1 corresponding to ≥500,000 somatic cells ml⁻¹ is commonly used as threshold of subclinical mastitis in temperate countries. However, given the innately high physiological level of somatic cells in low yielding dairy cows, this threshold may not apply to low yielding dairy cows. The current study was undertaken to investigate the clinical utility of CMT for screening of Staphylococcus aureus subclinical mastitis in low yielding smallholder dairy cows in Tanzania. A total of 1151 of quarter-milk samples were CMT tested, of these 914-originated from cows with a lactation period of 14–305 days. All samples were screened for subclinical mastitis by the CMT as well as microbiological culture of single, duplicate (two consecutive) and triplicate (three consecutive) samples as a gold standard. For the duplicate and triplicate quarter-samples, cows were considered positive for S. aureus subclinical mastitis if results of microbiologic culture for S. aureus were positive for two of two, and for at least two of the first three consecutive quarter-milk samples collected from that cow, respectively. Using a CMT score of ≥1 would classify 78.6% of the 940 quarter-samples as positive. Eighty-two percent of the samples in which S. aureus was isolated had CMT scores ≥2; this would classify 51.6% of the 940 quarter-samples as positive. For the single sample, this cut-off had sensitivity, specificity and likelihood ratio for S. aureus of 0.87, 0.83 and 4.24, respectively. For the duplicate quarter-milk samples this cut-off had sensitivity, specificity, and likelihood ratio of 0.94, 0.86, and 5.19. While, for the triplicate quarter-milk samples this cut-off had sensitivity, specificity and likelihood ratio of 0.97, 0.92 and 7.47, respectively. Based on these results and practical considerations, it is concluded that CMT score of ≥2 corresponding to ≥800,000 somatic cells Ml⁻¹ is the best cut-off to correctly identify S. aureus intramammary infections in low yielding dairy cows in Tanzania.     

Grass steer production system to improve carcass and meat quality

Three different production systems for autumn born dairy steers, managed more or less intensively, have been investigated. The aim of the production systems were to use a basic diet of grass, either grazed or conserved as silage or hay, combined in various proportions. The animals were slaughtered, aiming for the same degree of fattening, but at three ages. The intensive management aimed at slaughter in January–February at 26–28 months of age after indoor finishing with silage or hay. An “intensive out-of-season” management wished improve at slaughter at 28–30 months of age. Finally an extensive management with finishing at grazing resulted in slaughter in May–June at 30–33 months of age.

In three successive factorial experiments, 106 steers of two genotypes, Holstein and Montbéliard were reared to test the effects of the three production systems. The animals were evaluated for productive traits, carcass quality and biochemical characteristics of m. longissimus thoracis and m. rhomboideus thoracis.

For both breeds, the “extensive” animals produced heavier carcasses (+ 20 kg between “intensive” and “extensive”; P < 0.002). The carcass conformation evaluated by the EUROP classification did not differ between the production systems. The estimated percentage of adipose tissue of the carcass was significantly lower for the “extensive” steers (P < 0.05). The shear force measured on raw meat was not significantly affected by the production system, but after cooking, the shear force was higher for the extensive management as was the total collagen content. The haeminic iron content was always lowest in the intensive management. The results of these three series of experiments show that it is possible to produce steers with a feeding regime consisting mainly of grass (from 87% to 94% of the dry matter intake) grazed or harvested as hay or silage. To make the best use of the herbage resources of the farm and to limit the purchase of concentrates, the production of autumn born steers slaughtered at 32 months seems to be an attractive production system for dairy farmers.     

Herd-level risk factors for Salmonella enterica subsp. enterica in U.S. market pigs

Midwest U.S. herds (n = 63) were studied to identify risk factors for harboring Salmonella enterica among slaughter-weight pigs. Samples collected on farms (feces) and at slaughter (distal colonic content, cecal content and ileocolic lymph nodes) were cultured using conventional means. Approximately 15 pigs were studied per herd, for a total of 3754 samples. The proportion of pigs positive in one or more samples was calculated for each herd. Herd characteristics were described by a combination of interview and written survey. Logistic regression was used to detect relationships between the detection of Salmonella and potential herd-level risk factors. The mean individual pig prevalence was 5% for feces, 4% for distal colonic content, 15% for ileocolic lymph nodes, and 17% for cecal contents. One or more Salmonella isolates were detected in at least one sample type in every herd. The five most common serovars were S. Agona, S. Derby, S. Schwarzengrund, S. Typhimurium and S. Senftenberg, with 25 additional serovars detected. Salmonella prevalence estimates were positively correlated among all samples except distal colonic content and ileocolic lymph nodes. Pigs with culture positive fecal samples were at increased odds of being detected positive for each of the slaughter-collected samples examined, namely distal colonic content (OR = 30.5), ileocolic lymph nodes (OR = 12.9) and cecal content (OR = 23.2). Herds with positive fecal sample(s) had increased odds of having positive cecal content (OR > 1.5), distal colonic content (OR = 15.3) and ileocolic lymph nodes (OR = 12.7). Pigs from herds with at least some bowl drinkers had eight-fold higher odds of testing Salmonella positive than did pigs from herds with only nipple drinkers. Pigs from herds with only dry feeders had five-fold higher odds of testing Salmonella positive when compared with pigs from herds with combinations of wet/dry style feeders. Interventions at these two points should be considered when designing growing pig facilities to reduce Salmonella shedding.     

Management factors associated with Babesia bovis seroprevalence in cattle from eastern Yucatán, Mexico.

The effect of management on the seroprevalence of Babesia bovis was studied in 399 Bos indicus cattle 1–2 years old from 92 farms in the eastern Yucatán, México. The management factors studied were: farm-type, production system, herd size, farm size, stocking density, vector control, dipping interval, type of dipping, type of acaricide and cattle introduction to the farm. A cross-sectional study was carried out (2-stage cluster sampling). The number of serum samples was proportionally distributed according to the number of farms in the nine locations of eastern Yucatán, México (399 animals from 92 farms). Antibody activity to B. bovis was tested using an indirect ELISA. The farms with a seroprevalence ≤75% were considered as cases and those with seroprevalence >75% were considered as controls. The variables with p ≤ 0.20 were included in fixed effects logistic regression. The seroprevalence of the zone was 73.8% (66.3–81.3%). The following risk factors were found: Stocking density (<1 head/ha, OR = 4.04, CI (OR) = 1.20–13.62) and dipping interval (>60 days, OR = 5.07 CI (OR) = 1.26–20.48).     

Prevalence of agglutinating antibodies to Sarcocystis neurona in raccoons, Procyon lotor, from the United States.

Equine protozoal myeloencephalitis (EPM) is the most important protozoal disease of horses in North America and it is caused by Sarcocystis neurona. Natural cases of encephalitis due to S. neurona have been reported in raccoons, Procyon lotor. We examined 99 raccoons for agglutinating antibodies to S. neurona using the S. neurona agglutination test (SAT) employing formalin-fixed merozoites as antigen. Raccoons originated in Florida (N=24, collected in 1996), New Jersey (N=25, collected in 1993), Pennsylvania (N=25, collected in 1999), and Massachusetts (N=25, collected in 1993 and 1994). We found that 58 (58.6%) of the 99 raccoons were positive for antibodies to S. neurona using the SAT; 44 of 99 raccoons (44%) had titers of ≥1:500. This prevalence is similar to the reported seroprevalence of 33–60% for S. neurona antibodies in horses from the United States using the Western blot test.     

Transduction of bla(CMY-2), tet(A), and tet(B) from Salmonella enterica subspecies enterica serovar Heidelberg to S. Typhimurium

Antimicrobial resistance of Salmonella spp., especially resistance mediated by extended spectrum β-lactamases (ESBL), is a growing public health concern. Understanding the mechanisms through which Salmomella spp. acquire the resistance genes can lead to the development of intervention and mitigation strategies. Thirty-one Salmonella isolates of bovine origin were analyzed by serotyping, antimicrobial susceptibility testing, phage induction and bacterial host range determination, and phage transduction of antimicrobial resistance. The Salmonella isolates consisted of 12 serovars. Resistance to 1 or more antibiotics was detected in 12 isolates. Inducible phages were recovered from 19 Salmonella (61%) by spot lysis assay. Of the 19 phage samples, 13 were able to multiply in 2 or more Salmonella of various serovars. A cross-serovar transduction of antimicrobial resistance was observed from S. Heidelberg to S. Typhimurium. Transfection of an antimicrobial-susceptible strain of S. Typhimurium with a phage propagated in a S. Heidelberg resistant to multiple β-lactam antibiotics and tetracycline resulted in independent acquisition of bla_CMY-2, tet(A), and tet(B). These data indicate that inducible phages are common in Salmonella of bovine origin, many of them demonstrate a broad host range, and wild-type phage mediated transduction may contribute to the dissemination of antimicrobial resistance, including the resistance mediated by ESBL.     

Prevalence of intestinal pathogens in Danish finishing pig herds

Our aim was to determine the prevalence of the intestinal bacteria: Lawsonia intracellularis, Brachyspira hyodysenteriae, Serpulina intermedia, Brachyspira innocens, Brachyspira pilosicoli, pathogenic Escherichia coli (serogroups O138, O139, O141 and O149) and Salmonella enterica in Danish finishing pig herds. A total of 79 herds was randomly selected and visited during 1998. From each herd, 20 faecal samples were collected from individual pigs weighing 30–50 kg. Furthermore, 10 pooled pen samples were collected and examined for S. enterica. In total, 1580 faecal samples and 790 pen samples were collected and examined by polymerase chain reaction (PCR) or culture. L. intracellularis was found in 74 herds (93.7%), B. hyodysenteriae in two herds (2.5%), S. intermedia in 10 herds (12.7%), B. innocens in 27 herds (34.2%), B. pilosicoli in 15 herds (19.0%), pathogenic E. coli in 19 herds (24.1%) and S. enterica in eight herds (10.1%). The within-herd prevalences of L. intracellularis and B. hyodysenteriae were 25–30%; the within-herd prevalences of the other agents were 5–10%. Three herds (4%) were not infected with any of the bacteria and 25 herds (32%) were only infected with L. intracellularis.     

Financial analysis of various strategies for the control of Neospora caninum in dairy cattle in Switzerland

The present study was conducted to estimate the direct losses due to Neospora caninum in Swiss dairy cattle and to assess the costs and benefits of different potential control strategies. A Monte Carlo simulation spreadsheet module was developed to estimate the direct costs caused by N. caninum, with and without control strategies, and to estimate the costs of these control strategies in a financial analysis. The control strategies considered were “testing and culling of seropositive female cattle”, “discontinued breeding with offspring from seropositive cows”, “chemotherapeutical treatment of female offspring” and “vaccination of all female cattle”. Each parameter in the module that was considered to be uncertain, was described using probability distributions. The simulations were run with 20,000 iterations over a time period of 25 years.

The median annual losses due to N. caninum in the Swiss dairy cow population were estimated to be € 9.7 million. All control strategies that required yearly serological testing of all cattle in the population produced high costs and thus were not financially profitable. Among the other control strategies, two showed benefit–cost ratios (BCR) >1 and positive net present values (NPV): “Discontinued breeding with offspring from seropositive cows” (BCR = 1.29, NPV = € 25 million) and “chemotherapeutical treatment of all female offspring” (BCR = 2.95, NPV = € 59 million). In economic terms, the best control strategy currently available would therefore be “discontinued breeding with offspring from seropositive cows”.     

1株跨属感染猪霍乱沙门菌和大肠杆菌烈性噬菌体的分离及其生物学特性

从健康猪肠道内容物分离鉴定猪霍乱沙门菌（Salmonella choleraesuis,S.choleraesuis）的烈性噬菌体,并对其进行生物学特性分析。以S.choleraesuis 13122为宿主菌,用双层平板法从猪肠道黏膜和食糜样品中分离噬菌体,用电镜形态观察和基因组测序确定其分类,用点滴法和双层平板法测定其在大肠杆菌上的宿主谱,用浊度法测定其裂菌效力。分离到1株猪霍乱沙门菌噬菌体,命名为沙门菌噬菌体L6jm（Salmonella phage L6jm）。L6jm头部呈正多面体直径约75 nm,具有非收缩性的尾部,长约190 nm,其基因组无整合酶基因和细菌基因组,故判定为烈性噬菌体;L6jm可感染S.choleraesuis 13122并跨属感染1株大肠杆菌（Escherichia coli,E.coli）244,同时,可裂解9株大肠杆菌,包括1株产肠毒素大肠杆菌（ETEC）;L6jm在S.choleraesuis 13122和E.coli 244上的最佳感染复数分别为0.01和0.001;潜伏期均为15 min,暴发期分别为145和125 min;L6jm在≤50℃,pH为3~11时稳定;L6jm在液体培养基中对S.choleraesuis 13122有显著（P<0.05）的抑菌效果,对ETEC104在MOI 100时也可产生显著（P<0.05）的抑菌效果。L6jm可跨属感染S.choleraesuis和E.coli,且能裂解1株ETEC,具有防治沙门菌和大肠杆菌感染的应用潜力。     

Cryptosporidiumparvum oocysts in seawater clams (Chameleagallina) in Italy

Bivalves filter large volumes of water and can concentrate organisms which are pathogenic for humans and animals. Our aim was to evaluate the presence of Cryptosporidium spp. in clams from the Adriatic coast (Abruzzo region) and genetically characterize the oocysts isolated from the clams. From March to July 2003, 960 specimens of clams (Chamelea gallina) present in nature were collected at 500 m from the Tordino, Tronto, Vibrata and Vomano river mouths on the Adriatic sea. The haemolymph and tissues were extracted from the specimens (240 per river mouth) after the specimens had been identified, measured and weighed (live weight). Immunofluorescence tests (IFA) were performed on pools (n = 32) of samples and oocysts of Cryptosporidium spp. were detected in 23 pools of C. gallina. To identify the Cryptosporidium species, all the pools IFA-positive were tested by a PCR assay specific for the Cryptosporidium outer wall protein (COWP) gene. Positive amplicons then were sequenced and analysed. Two pools of clams were positive for Cryptosporidium parvum Genotype 2 (the “bovine” i.e. zoonotic genotype). This is the first time that C. parvum was found in clams from the Adriatic sea in Italy and the case might be of public health importance.     

Genomic lineage of Salmonella enterica serovar Dublin

Thirty five strains of the host adapted Salmonella serotype Dublin (S. Dublin) have been characterized by IS200 patterns, ribotyping, pulsed-field gel electrophoresis (PFGE), restriction fragment polymorphism after hybridization with five randomly cloned DNA-framents of S. enteridis (RFLP), and plasmid profiling in order to divide the strains into ‘genomic lines’. For comparison, 20 other strains of 9 different group-D serotypes were included. The IS200 patterns were identical in all strains of S. Dublin examined. These patterns were different from those observed in other group-D Salmonella with the exception of one strain S. Enteritidis phage type 11 and a strain of S. Rostock. The insertion element IS200 was not detected in strains of S. Dar-es-Salam, S. (II) 9,12:z -, and S. Panama. RFLP, based on probing with five random cloned chromosomal fragments gave the same pattern in all strains except for one isolate from the UK. This strain was also found to have an unique PFGE pattern and ribotype. Among the remaining strains, three different PFGE patterns and 7 different ribotypes were observed. Based on all four typing methods, 8 different ‘genomic lines’ of S. Dublin were identified. The same grouping could be obtained from the use of ribotyping alone, but PFGE and RFLP were found to provide valuable information on possible relationships between ribotypes. Seven different plasmid profiles and a group of strains without plasmids were observed. In several cases, the same plasmid profile was shown to be present in more than one ‘genomic line’.     

牦牛与普通牛、水牛的X染色体基因编码区比较及密码子偏性分析

【目的】 从基因组比对及密码子偏性角度分析牦牛与其他牛亚科动物X染色体,有助于了解牦牛品种差异及系统进化地位,为其适应高原低压低氧环境和密码子优化提供参考。【方法】 以牦牛X染色体参考基因编码区序列为参考,与普通牛和江河型水牛X染色体参考基因编码区序列进行基因组比对和基因共线性分析,同时进行基因注释,对过滤后的编码区文件进行相对同义密码子使用频率（RSCU）、ENC-plot、PR2-plot和最优密码子确定等偏性分析。【结果】 在牦牛X染色体基因编码区发现了参与气管收缩、肺部呼吸及机体代谢等基因与普通牛和水牛存在差异,如KLHL13、CENPI和PGK1等基因;共线性显示长段由强选择压和突变压下表现出的交换线性区域;密码子分析中3种牛亚科动物密码子使用偏性相似,偏性均较弱,均偏向G/C结尾;强偏性密码子（RSCU≥1.5）均为CUG、GUG、AGA、AGG和UGA;牦牛、普通牛和水牛分别筛选出16、13和9个最优密码子,均以A/U结尾。【结论】 牦牛与普通牛、水牛相比面临了更大的选择压力,累计的变异程度更大,三者的密码子偏性受到自然选择作用均大于突变作用。研究结果为牦牛的遗传育种、密码子优化和遗传资源开发利用提供参考。     

Efficacy of spheroplastic and cell-wall competent vaccines for Mycobacterium avium subsp. paratuberculosis in experimentally-challenged baby goats

A Mycobacterium avium subspecies paratuberculosis (MAP) vaccine that reduced the incidence of clinical disease or reduced fecal shedding of MAP would aid control of Johne's disease (JD). The objective of the present study was to evaluate the efficacy of four MAP vaccine combinations, including cell-wall competent (CWC) alum adjuvant, CWC-QS21 adjuvant, cell-wall deficient (CWD) alum adjuvant and CWD-QS21 adjuvant vaccines. Eighty baby goats were vaccinated at 1 and 4 weeks of age with one of these vaccines or a sham control vaccine consisting of alum adjuvant. Kids were challenged orally with approximately 6.0 × 10⁹ organisms in four divided doses of 1.5 × 10⁹ organisms using a goat isolate of MAP. Vaccinated challenged and challenged control groups had 10 and 6 kids per group, respectively. Half of the kids within each group were necropsied at either 6 or 9 months post-challenge. Gross and microscopic lesions and relative number of acid-fast bacilli were evaluated and scored at necropsy. Results indicated all challenged kids had some lesions compatible with JD suggesting none of the vaccines prevented infection. Three vaccines (CWC-alum, CWC-QS21 and CWD-QS21) reduced lesion scores by 46–51% at 9 months. CWD-alum vaccine resulted in a more severe (+33.5%) lesion score than sham-vaccinated challenged control. Lesion scores were greater at 9 months than at 6 months post-challenge in the sham-vaccinated challenged group and CWD-alum vaccinated group, while lesion scores were generally stable with remaining vaccines. Mean fecal CFU/g were significantly different across time from challenge to 9-month necropsy (p = 0.043) and the CWC-QS21 vaccine group had a marked reduction in fecal CFU/g at all time points post-challenge. A reduction in MAP CFU/g was also detected in necropsy tissues from kids given the CWC-alum, CWC-QS21 and CWD-QS21 vaccines, and increased CFU/g were detected in tissues from kids given the CWD-alum vaccine. Immunological tests evaluated included, humoral response evaluation by AGID, ELISA and Western blot, and cell mediated response by comparative PPD skin testing (M. avium, Old Johnin, M. bovis and Lot 2 Johnin PPD's), and production of MAP induced γ-interferon. Vaccination also resulted in false-positive PPD skin test reactions for M. avium PPD, Old Johnin PPD and γ-interferon tests. When a 2-mm cutoff above normal skin thickness was used to define positive skin test reactions, false-positive reactions for M. bovis were detected in only 2 of 32 kids given a vaccine with QS21 adjuvant.