Endotoxin in the conscious piglet: Its effects on some general and gastrointestinal myoelectrical parameters

The effect of an intravenous bolus injection of endotoxin, 0.1, 1 or 10 g/kg, on rectal temperature, clinical appearance, haematological parameters, and on gastrointestinal electrical activity was examined in 11 conscious piglets of 4–5 weeks of age, with implanted electrodes in the antrum pylori, duodenum, jejunum and ileum. All doses resulted in a significant and dose-dependent increase in rectal temperature, in pronounced clinical signs and in distinct changes in haematological values. These included shivering, depression, respiratory distress, a leukopenia (0.1 g/kg) or a leukocytosis (1 g/kg) with a shift to the left, an accelerated sedimentation rate and a decreased packed cell volume. Doses of 1 and 10 g/kg induced a transient inhibition of gastroduodenal electrical activity. These results suggest that, in the piglet, endotoxin primarily manifests general clinical signs and that the gastrointestinal effects coincide with these.     

The influence of anti-inflammatory therapy on bacterial clearance following intramammary Escherichia coli challenge in goats

Coliform mastitis in dairy cattle frequently results in systemic disease with occasional deaths in association with endotoxic shock. Systemic anti-inflammatory therapy has been used to alter the course of endotoxic shock in severe cases. Use of anti-inflammatory therapy has been questioned on the basis that such treatment may compromise immune function and decrease clearance of bacteria from infected mammary glands. The purpose of this investigation was to determine whether anti-inflammatory therapy influenced bacterial clearance following intramammary challenge of lactating goats with Escherichia coli.Standardized quantities of a pathogenic coliform culture were infused through the teat canal into one half of the mammary gland in 18 goat does. The does were then randomly assigned to receive one of three intravenous treatments: saline (controls), one dose of steroid (dexamethasone), or two doses of a non-steroidal anti-inflammatory agent (flunixin meglumine). The clinical signs, milk production, complete blood counts, serum clinical chemistry values, milk bacterial cultures and milk somatic cell concentrations were monitored sequentially.Goats treated with anti-inflammatory agents exhibited some improvement in clinical response to challenge with E. coli (e.g. rectal temperature, degree of appetite suppression) as compared to saline controls. There were no significant differences between treatments in the degree of inflammation present in the mammary glands or supramammary lymph nodes examined at necropsy. The most important finding was that anti-inflammatory therapy did not adversely influence the clearance of E. coli from challenged glands.     

Inhibitory effects of SKF96365 on the activities of K+ channels in mouse small intestinal smooth muscle cells

In order to investigate the effects of {"type":"entrez-protein","attrs":{"text":"SKF96365","term_id":"1156357400","term_text":"SKF96365"}}SKF96365 (SKF), which is a non-selective cationic channel blocker, on K⁺ channel currents, we recorded currents through ATP sensitive K⁺ (I_KATP), voltage-gated K⁺ (I_Kv) and Ca²⁺ activated K⁺ channels (I_BK) in the absence and presence of SKF in single small intestinal myocytes of mice with patch-clamp techniques. SKF (10 µM) reversibly abolished I_KATP that was induced by cromakalim (10 µM), which is a selective ATP sensitive K⁺ channel opener. These inhibitory effects were induced in a concentration-dependent and voltage-independent manner. The 50% inhibitory concentration (IC₅₀) was 0.85 µM, which was obviously lower than that reported for the muscarinic cationic current. In addition, SKF (1 µM ≈ the IC₅₀ value in I_KATP suppression) reversibly inhibited the I_Kv that was induced by repetitive depolarizing pulses from −80 to 20 mV. However, the extent of the inhibitory effects was only ~30%. In contrast, SKF (1 µM) had no significant effects on spontaneous transient I_BK and caffeine-induced I_BK. These results indicated that SKF inhibited ATP sensitive K⁺ channels and voltage-gated K⁺ channels, with the ATP sensitive K⁺ channels being more sensitive than the voltage-gated K⁺ channels. These inhibitory effects on K⁺ channels should be considered when SKF is used as a cationic channel blocker.     

Protective effect of CpG-DNA against mastitis induced by Escherichia coli infection in a rat model

A mastitis model in rats, induced by Escherichia coli infection, was established and the protective effect of Cytosine-phosphate-Guanosine (CpG)-DNA was determined. An E. coli suspension containing either 2 x 10(3) colony forming units (CFU)mL(-1)(EL group), 2 x 10(5)CFU mL(-1) (EH group), or (as controls) 100 microL phosphate buffer saline (CON group), was inoculated into the mammary glands 72 h after parturition. The rats were euthanased 24 h post-infection. The histopathological changes in mammary tissue in the EL group were mild, whereas the structural changes in the EH group were severe and polymorphonuclear leukocytes (PMNs) had accumulated in the mammary alveoli. Interleukin (IL)-6 and tumour necrosis factor (TNF)-alpha and N-acetyl-beta-d-glucosaminidase (NAGase) were significantly increased in the mammary tissue from the EH group but not significantly changed in the EL group. On the basis of these findings, the potential protective effect of CpG-DNA on mammary glands was tested using a 2 x 10(5)CFU mL(-1) suspension. An intramuscular injection of either CpG-DNA (200 microg) or PBS (100 microL) was given immediately after parturition. At 72 h post-partum, 2 x 10(5)CFU mL(-1)E. coli (100 microL) were inoculated into the mammary glands of all rats. At pre-infection (0 h), and 8, 16, 24, 48 and 72 h after inoculation six rats were euthanased. CpG-DNA induced more rapid migration of PMNs from the blood to mammary tissue at the initial stage of infection, stimulated the secretion of IL-6 and TNF-alpha at different time points, reduced viable E. coli in mammary tissues and decreased the activity of NAGase. CpG-DNA also promoted the expression of its specific receptor TLR-9 mRNA in mammary tissue. The study showed that CpG-DNA protected against E. coli mastitis in this rat model.     

The effects of experimentally induced fever on the estimated blood flow to and oxygen utilization by the liver and the viscera drained by the portal vein in sheep

Intrajugular injection of a purified E. coli lipopolysaccharide induced a biphasic fever in sheep after a latent period of 12 to 20 min. The changes in the blood flow from the liver and from the viscera drained by the portal vein were: (a) in the latent period, decreases in total hepatic blood flow (THF) due to decreased portal venous blood flow (PVF); (b) during the first febrile phase, increases in THF due to increased hepatic arterial blood flow and, (c) in the second febrile phase, increases in THF due to decreased PVF. Although there were large variations in the oxygen supply to the viscera drained by the portal vein and to the liver, there were relatively small or no changes in their oxygen consumption.     

Field trial evaluating changes in prevalence and patterns of antimicrobial resistance among Escherichia coli and Enterococcus spp. isolated from growing broilers medicated with enrofloxacin, apramycin and amoxicillin

The present study investigates, under field conditions, the influence of antimicrobial administration on prevalence and patterns of antimicrobial resistance among Escherichia coli and Enterococcus spp. isolated from growing broilers. For this purpose, a group of 16,000 commercial broiler chickens was treated with enrofloxacin from day 1 to day 3, gentamicin from day 19 to day 21, and ampicillin from day 26 to day 28. A control group of 16,000 broilers was placed in the same controlled environment poultry house. Fecal (from both groups) and feed samples were collected at regular intervals. Few E. coli isolates were obtained from either farm environment or poultry feed samples, while enterococci were found to be ubiquitous among these samples. The frequency of resistance against most antimicrobials tested was significantly higher (P < 0.05) in E. coli isolated from broilers receiving intermittent antimicrobial pressure than that from non-medicated broilers, whereas in enterococci these differences were only observed among structurally related antimicrobial drugs and over a short period of time. By the time the broilers reached market age (33 days), several multi-resistant E. coli and enterococci were detected in the feces of the medicated group. Results suggest that antimicrobial resistance in E. coli was mainly medication-dependent, whereas among enterococci, changes observed over time were apparently influenced by factors apart from antimicrobial exposure, namely the resistance organisms previously present in farm environment and those present in feedstuffs.     

Verocytotoxins (Shiga-like toxins) produced by Escherichia coli: a minireview of their classification, clinical presentations and management of a heterogeneous family of cytotoxins

Bacterial virulence usually requires the interaction of multiple factors in order to cause disease. The enterotoxins produced by certain strains of bacteria are proteins which vary in their mode of action, but do fall into two general groups; the cytotoxic and the cytotonic enterotoxins. While cytotoxic enterotoxins typically kill eucaryotic cells (eg. by inhibiting protein synthesis), cytotonic enterotoxins derange cell metabolism in specific ways (eg. by elevating cyclic nucleotide levels). Some strains of Escherichia coli produce protein toxins that are biologically, structurally and antigenically related to a cytotoxin (Shiga toxin) (ShT) produced by Shigella dysenteriae type 1. Although this group of related, but not necessarily identical toxins have been referred to as Vero cell toxins or Verocytotoxins (VTs), the term Shiga-like toxins (SLTs) has been widely accepted. ShT and SLTs have been implicated as a cause of diarrhoea as well as haemorrhagic colitis (HC) and haemolytic uremic syndrome (HUS) in humans, whilst SLTs have been implicated as causal agents of oedema disease and HC in weaner pigs and calves, respectively. While S. dysenteriae is an invasive organism, the SLT-producing strains of E. coli have not been reported to be invasive, but cause diarrhoea that may contain blood and mucus. Thus, SLTs can be considered an important "new" type of enterotoxins whose role in the pathogenesis of diarrhoea, HC and HUS is beginning to emerge, not only in certain geographical settings, but worldwide. This mini review focuses on this family of SLTs, because of recent advances which have been made towards their detection, nomenclature, pathogenesis and possible management of their clinical presentations.     

Effect of susceptibility to enterotoxigenic Escherichia coli F4 and of dietary tryptophan on gut microbiota diversity observed in healthy young pigs

Healthy weaned pigs susceptible to enterotoxigenic Escherichia coli F4 (ETEC) require more tryptophan (Trp) to maximize their performance. This may be related to an effect on intestinal microbiota. We studied the intestinal bacterial diversity of healthy pigs with different susceptibility to ETEC and fed different Trp levels. Thirty-six littermate weaned pigs were selected to obtain a set potentially formed of 50% ETEC-susceptible and 50% non-susceptible pigs, based on a Mucin 4 gene polymorphism. Pigs were fed a diet with 0.17 (TrpL) or 0.22 (TrpH) standardized ileal digestible Trp:Lys ratio for 21 days. Slaughtered pigs were classified into non-susceptible, mildly susceptible, and susceptible, by testing ETEC adhesion to intestinal villi. Bacterial diversity in jejunum content was assessed by the 16S rRNA gene-targeted denaturing gradient gel electrophoresis (DGGE) fingerprinting analysis and expressed by the Shannon index. Susceptible pigs had a reduced bacterial diversity, particularly with TrpL diet (p = 0.003). The ETEC adhesion class affected the quantification of enterobacteria DNA (p = 0.027). One DGGE band, which referred to Clostridium bartlettii, was not evidenced in all the susceptible pigs; less DNA from this microbe was quantified by RT-PCR in the jejunum from TrpH susceptible pigs (p = 0.025) compared to TrpL. The gene expression for β-galactoside α-2,3-sialyltransferase 1 was higher in jejunal tissue of ETEC-susceptible pigs (p = 0.019). In studies on pig gut microbiota, the presence of intestinal receptors for ETEC should be considered because of their contribution to a reduced bacterial diversity. This effect could be partially reversed by dietary Trp addition.     

Influence of dietary condensed tannins from sericea lespedeza on bacterial loads in gastrointestinal tracts of meat goats

This research assessed the potential use of a low input forage containing a high amount of condensed tannins (CT) to reduce foodborne pathogens prior to slaughter of meat goats. In a completely randomized design, twenty Kiko × Spanish intact male kids (BW = 19.2 ± 0.74 kg) were fed ground sericea lespedeza [SL; Lespedeza cuneata (Dum-Cours) G. Don; 2 pens], a high-CT legume, or bermudagrass hay [BG; Cynodon dactyon (L.) Pers.; 2 pens], at 75% of daily intake with a corn-based supplement (25% of intake) for 14 weeks (n = 10 goats/treatment). At the end of the feeding trial, the animals were slaughtered using standard procedures. Immediately after evisceration, rumen and rectal samples were collected to assess bacterial loads and volatile fatty acids in the rumen. Concentrations of rumen volatile fatty acids were significantly different between dietary treatments. Goats fed SL hay had higher (P < 0.05) contents of butyric (8.66 vs 7.16 mM), isobutyric (1.94 vs 1.44 mM), isovaleric (3.03 vs 2.13 mM), and valeric (1.43 vs 1.07 mM) acids than those fed BG hay; however, the content of acetic acid (78.6 vs 64.4 mM) was higher (P < 0.05) in the BG-fed groups than in SL-fed groups. Escherichia coli (2.33 vs 1.13 log₁₀ CFU/g) counts of rumen contents were higher (P < 0.05) in the SL-fed group compared with the BG-fed group. However, E. coli counts in feces were not different (P > 0.05) between dietary treatments. The high-CT influenced (P < 0.05) total plate counts in the feces; and the total plate counts in feces of SL- and BG-fed goats were 4.95 and 6.57 log₁₀ CFU/g, respectively. The results indicated that high CT in the diet might influence rumen volatile fatty acid composition, but might not reduce the bacterial loads in gastrointestinal tracts of meat goats.     

Effect of antisecretory drugs on experimentally induced weanling diarrhoea in piglets

In 45 newly-weaned 3 to 4-week-old piglets, diarrhoea was induced by a combined infection with transmissible gastroenteritis (TGE) virus and enterotoxigenic E. coli (ETEC) strains. In untreated control animals this dual inoculation resulted in profuse diarrhoea, vomiting, hypovolaemic shock and death of 77% of the animals within five days of TGE virus inoculation. Antisecretory drugs were administered intramuscularly for three consecutive days after experimental infection. The neurolepticum chlorpromazine, at 2 mg/kg/24 h, resulted in a significant inhibition of diarrhoea and vomiting, and in an increase in weight gain and survival. Sedation and hypothermia, however, were serious side-effects. The ₂ agonist clonidine, at 80 g/kg/12 h, induced a significant antidiarrhoeal effect and a reduction in mortality. The drug, however, provoked decreased activity of ₂-adrenergic excitation and incoordination. The -adrenergic antagonist propranolol, at 0.33 mg/kg/8 h, and the calcium channel blocker verapamil, at 2 mg/kg/8 h, had no beneficial effect on the experimentally induced diarrhoea.     

川西北牦牛肉中产志贺毒素大肠杆菌的分离鉴定及stx2基因的序列分析

为了解中国川西北牦牛肉中产志贺毒素大肠杆菌(STEC)的携带情况及stx2的亚型和特征,试验将采集的204份川西北牦牛肉样品(各25g)增菌培养后,每份挑取5个可疑菌落,采用stx1、stx2双重PCR方法检测STEC,对分离株中的stx2分型并克隆测定stx2编码区序列。结果显示,在204份样品中分离出8株STEC,平均分离率为3.9%(8/204);存在4个不同的O血清型,分别为O38(4)、O50(1)、O74(2)、O150(1);在6株含有stx2的菌株中,其中有2株为stx2a型、4株为stx2c型。结果表明牦牛源分离株氨基酸序列与人源和牛源菌株同源性较高;由stx2A、B亚基的氨基酸序列系统进化树可知,牦牛源分离株与人源、牛源菌株聚为一支,表明它们之间遗传距离相对较近,牦牛源stx2各自分布在自己的小分支中,表明牦牛源STEC stx2与人源和牛源stx2相比,尽管亲缘关系较近,但仍存在一定程度的差异。     

Effect of bacitracin on tetracycline resistance in Escherichia coli and Salmonella

Tetracyclines and bacitracin are used extensively as a growth promotant and a therapeutic, respectively in livestock. In this study, we test whether there is an interaction between bacitracin and tetracycline that can select for an increase in microbial tetracycline sensitivity, using Escherichia coli and Salmonella as a model. We found via in vitro studies that bacitracin at sublethal concentrations preferentially selects for outgrowth of tetracycline-sensitive bacteria from among a population of tetracycline-resistant and tetracycline-sensitive E. coli and Salmonella strains. Most of the bacterial strains employed in our study were shown by PCR to possess the tetracycline resistance gene tet(A) or tet(C), either of whose activity is associated with tetracycline efflux. Bacitracin could be substituted for the lipophylic chelator, fusaric acid, used as a positive selective agent for tetracycline sensitivity. Together, these observations suggest that bacitracin-mediated selection for tetracycline sensitivity alters the function of tetracycline efflux proteins. Using bacitracin and tetracycline simultaneously may improve the effectiveness of the antibiotics, while decreasing the risk of selecting for a population of tetracycline-resistant microorganisms.     

Prevalences of resistance to seven antimicrobials among fecal Escherichia coli of swine on thirty-four farrow-to-finish farms in Ontario, Canada

Fecal specimens were composited and a hydrophobic-grid membrane-filter method was used to measure antimicrobial resistance to ampicillin 16 μg/ml, carbadox 30 μg/ml, gentamicin 4 μ/ml, nitrofurantoin 32 μg/ml, spectinomycin 16 μg/ml, sulfisoxazole 32 μg/ml and tetracycline 8 μg/ml among 8119 Escherichia coli isolates from 68 fecal samples collected on 34 farrow-to-finish swine farms marketing over 500 hogs/yr. The overall prevalences of resistance to antimicrobials among these isolates were: ampicillin 29%, carbadox 3.5%, gentamicin 0.6%, nitrofurantoin 27%, spectinomycin 28%, sulfasoxizole 38% and tetracycline 71%. Thirty to seventy-six per cent of the variations in prevalences were explained by between-farm differences.     

猪大肠杆菌和葡萄球菌的分离鉴定及耐药性分析

对湘西北地区某猪场患病的仔猪的粪便及病变的脾脏、肝脏、淋巴结、肺脏、肾水肿液进行病原菌分离鉴定,并用Kirby-Bauer法测定分离菌株对15种抗菌药的敏感性。结果表明,患病仔猪分离到大肠杆菌和猪葡萄球菌两种细菌。其中大肠杆菌对庆大霉素、头孢呋肟、丁胺卡那霉素、四环素、先锋必素敏感,对氯霉素、氧哌嗪青霉素、先锋五号、头孢曲松钠、新霉素中介,对羧苄青霉素、强力霉素、青霉素、氨苄青霉素、红霉素有较高的耐药性。葡萄球菌对先锋霉素Ⅴ、丁胺卡那霉素敏感,对头孢曲松钠中介,对其他抗生素耐药。其中大肠杆菌对药物敏感率、中介率、耐药率均为33.33%;葡萄球菌的敏感率为13.33%,中介率为6.67%,耐药率为80.00%。     

The food contaminant fumonisin B1 reduces the maturation of porcine CD11R1+ intestinal antigen presenting cells and antigen-specific immune responses, leading to a prolonged intestinal ETEC infection

Consumption of food or feed contaminated with fumonisin B₁ (FB₁), a mycotoxin produced by Fusarium verticillioides, can lead to disease in humans and animals. The present study was conducted to examine the effect of FB₁ intake on the intestinal immune system. Piglets were used as a target and as a model species for humans since their gastro-intestinal tract is very similar. The animals were orally exposed to a low dose of FB₁ (1 mg/kg body weight FB₁) for 10 days which did not result in clinical signs. However, when compared to non-exposed animals, FB₁-exposed animals showed a longer shedding of F4⁺ enterotoxigenic Escherichia coli (ETEC) following infection and a lower induction of the antigen-specific immune response following oral immunization. Further analyses to elucidate the mechanisms behind these observations revealed a reduced intestinal expression of IL-12p40, an impaired function of intestinal antigen presenting cells (APC), with decreased upregulation of Major Histocompatibility Complex Class II molecule (MHC-II) and reduced T cell stimulatory capacity upon stimulation. Taken together, these results indicate an FB₁-mediated reduction of in vivo APC maturation.     

Epidemiological survey on <Emphasis Type="Italic">Escherichia coli</Emphasis> O157 in Chongqing and Three-Gorge Reservoir Areas of China

Prevalence, presence of virulence and adherence associated genes, genetic diversity, biochemical characteristics, and antibiotic susceptibility were determined for Escherichia coli O157 isolated over 4 months in Chongqing city and Three-Gorge Reservoir Areas. 11 isolates of E. coli O157 were isolated from 1504 samples and 7 of them are O157:H7 and 4 are O157:H? All O157:H7 isolates had eaeA, ehxA, EspA and Tccp genes, but did not have stx1 and stx2. All O157:H? isolates did not have stx1, stx2, eaeA, ehxA, EspA and Tccp genes except for the isolate obtained from Yunyang county which had stx1. When eaeA and ehxA presented in isolates were digested by restriction enzymes, the numbers and the sizes of the segments were the same as the control E. coli O157 strains. This suggests that eaeA and ehxA exhibit poor polymorphism. Most E. coli O157 isolates showed identical biochemical activities to the standard strains for sorbitol and rhamnose, and all E. coli O157:H7 obtained from feces at the same dairy cattle farm had similar biochemical characteristics. Antibiotic susceptibility demonstrated resistance of the isolates to penicillin, ampicillin, bacitracin, cefuroxime, erythromycin, gentamycin and tetracycline, indicating the isolates obtained in this study had a multi-drug resistance.     

Prevalence and age-related infection of Cryptosporidium suis, C. muris and Cryptosporidium pig genotype II in pigs on a farm complex in the Czech Republic

A total of 413 pig faecal samples were collected from pre-weaners (119), starters (131), pre-growers (123) and sows (40) from a farm with a closed breeding system segmented into two breeding complexes and a growing complex in the region of Vysočina, Czech Republic and screened for the presence of Cryptosporidium using staining methods and genotyping (SSU rRNA). Cryptosporidium oocysts were detected by microscopy in the faeces of 21.1% of the samples (87/413). Sequence analyses and RFLP identified C. suis in 44, Cryptosporidium pig genotype II in 23 and C. muris in 2 samples. No mixed infections were found.Pigs under 7 weeks of age were infected with C. suis only. Cryptosporidium pig genotype II was found in animals from 7 weeks of age. No relationship was found between diarrhoea and any Cryptosporidium infection in any of the different age groups (P < 0.05). The pre-weaned pigs shed significantly more Cryptosporidium oocysts than older pigs and it was associated with C. suis infection.     

Therapeutic effect of anti-TNF-α antibody and levofloxacin (LVFX) in a mouse model of enterohemorrhagic Escherichia coli O157 infection

The ability of an anti-TNF-α antibody to confer protection against enterohaemorrhagic Escherichia coli (EHEC) O157 was investigated in germfree IQI mice. The use of an antibiotic levofloxacin (LVFX) alone or with the antibody was also studied. Protection included an increase in survival rate. Treatment with the anti-TNF-α antibody inhibited the histological signs associated with EHEC infection but did not prevent the colonization of EHEC or production of Shiga toxin (Stx). No clinical signs were observed and EHEC was completely eliminated in the mouse model receiving both anti-TNF-α antibody and LVFX. Anti-TNF-α antibody suppressed inflammatory cytokine response in the mouse kidney and brain by EHEC infection.     

Construction,Expression, Purification,Refold and Activity Assay of a Specific ScFv Fragment against Foot and Mouth Disease Virus

An active form of a single-chain antibody (scFv) from the murine monoclonal antibody (mAb) 1C7, which is specific for type O foot and mouth disease virus (FMDV), was produced in Escherichia coli. The complementary DNAs encoding the variable regions of the heavy chain (V_H) and light chain (V_L) were connected by a (Gly₄Ser)₃ linker, using an assembly polymerase chain reaction. V_H-(Gly₄Ser)₃-V_L genes were screened by phage display technology. The sequencing results showed that the V_H gene of scFv was composed of germline VH76-1BG-DFL16.1-JH4 and the V_L gene of scFv consisted of germline bw20-JK2. The resultant scFv gene was cloned to the pProEX^TM HTc vector and expressed in E. coli as inclusion bodies. After extraction from the E. coli cells, the inclusion bodies were solubilized and denatured in the presence of 8 mol/L urea. The expressed scFv fusion proteins were purified by nickel–nitrilotriacetic acid and finally renatured by dialysis. The purity and activity of the purified scFv were confirmed by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and enzyme-linked immunosorbent assay. The result revealed that the 1C7 scFv conserved the same characteristics of specific recognition and binding to type O FMDV as the parental 1C7 mAb.