The Fecal and Serum Metabolomics of Giant Pandas Based on Untargeted Metabolomics.

Little is comprehensively known or understood about giant panda fecal and serum metabolites, which could serve as important indicators of the physiological metabolism of giant pandas. Therefore, we determined the contents of fecal and serum metabolites of giant pandas based on an untargeted metabolome. Four hundred and 955 metabolites were detected in the feces and serum of giant panda, respectively. Glycerophospholipid and choline metabolism were the main metabolic pathways in feces and serum. A significant correlation between the gut microbiota and fecal metabolites was found (P < 0.01). Fecal metabolites were not greatly affected by the age or gender of giant pandas, but serum metabolites were significantly affected by age and gender. The majority of different metabolites caused by age were higher in serum of younger giant pandas, including fatty acids, lipids, metabolites of bile acids, and intermediate products of vitamin D3. The majority of different metabolites caused by gender included fatty acids, phosphatidylcholine (PC), phosphatidylserine (PS), and phosphatidylethanolamine (PE). A separate feeding diet should be considered according to different ages and genders of giant panda. Therefore, our results could provide helpful suggestions to further protect captive giant pandas.

Dissipation and residue determination of penicillin G and its two metabolites in citrus under field conditions by DSPE/UPLC-MS/MS.

A rapid determination method of residual penicillin G and its two metabolites in citrus was developed and validated by dispersive solid-phase extraction and ultra-high performance liquid chromatography-tandem mass spectrometry (DSPE/UPLC-MS/MS). The samples were extracted with 80% acetonitrile and purified with octadecylsilane. High linearity was obtained with correlation coefficients (r2 ) >0.9981. The limits of quantification were 0.005-0.01 mg/kg. The recoveries of penicillin G and its metabolites spiked in blank citrus were within 76.7-107%, with relative standard deviations of 1.3-9.6%. The dissipation dynamics and distribution of penicillin G in citrus followed first-order kinetics, with half-life of 1.7-2.7 days. Penicillin G degraded easily in citrus and the metabolite was mainly penilloic acid, which can exist stably for long time. The terminal residues of penicillin G in pulp, whole citrus and peels were 0.015-0.701, 0.047-7.653 and 0.162-13.376 mg/kg, respectively. The hazard indexes for risk assessment of citrus were significantly <1, suggesting that the health risks to humans after consumption of citrus were insignificant and negligible. These results could provide necessary data for evaluating the safe and proper use of penicillin G in citrus.

Radiolabeling of benzylpenicillin with lutetium-177: Quality control and biodistribution study to develop theranostic infection imaging agent.

Benzylpenicillin acts through binding with beta-lactamase enzyme and inhibiting the bacterial cell wall biosynthesis. Therefore, the radiolabeling of benzylpenicillin with lutetium-177 is expected to serve as a theranostic agent for deep-seated bacterial infections. The radiolabeling of benzylpenicillin resulted ~93% radiochemical yield at optimized reaction conditions. Radiochemical purity analysis was tested with the help of Whatman No. 2 paper and instant thin layer chromatography. Biodistribution study with healthy New Zeeland white rabbit revealed moderate accumulation in different organs. Kidneys are the major organs, showed not more than 4.57±0.89% injected dose per gram organ (ID/gm organ) at 1 h time point and 3.48±1.11% ID/gm organ at 6 h time point. The accumulation of tracer agent in liver was found in the range of 7.42±2.42% to 9.09±2.76 ID/gm organ. The glomerular filtration rate studies revealed rapid clearance - omitting the chance of nephrotoxicity. The radiolabeling yield, biodistribution and glomerular filtration rate results revealed 177Lu-benzylpencillin could be a potential candidate to diagnose the deep-seated bacterial infection.

Evaluation of the efficacy of intramuscular versus intramammary treatment of subclinical Streptococcus agalactiae mastitis in dairy cows in Colombia.

A randomized controlled trial was performed in 17 Colombian dairy herds to determine the cure risk among cows subclinically infected with Streptococcus agalactiae exposed to 2 antibiotic therapies. Composite milk samples were collected before milking at the onset of the trial (pretreatment) and 2 subsequent times over a period of approximately 63 d. The intramammary application (IMM) of ampicillin-cloxacillin was compared with the intramuscular application (IM) of penethamate hydriodide, and cure risks after an initial and retreatment application were assessed. Cure risk after the initial treatment was higher (82.4%) for the IMM treatment than for IM therapy (65.8%). However, no difference was observed in the cure risk of refractory cases after retreatment (IMM=52.6% vs. IM=51.2%). The cumulative cure risk (both initial and retreatment) was 90.4 and 82.9% for the IMM and IM products, respectively. A 2-level random effects logistic model that controlled for pretreatment cow-level somatic cell count, indicated that IM treatment (odds ratio=0.37) had a lower cure risk than IMM and a tendency for a lower cure risk with increasing baseline somatic cell count. Our findings suggest that both products and administration routes can reduce the prevalence of S. agalactiae in affected herds, but the IMM product had a better efficacy in curing the infection. In addition to the treatment protocol, the cow somatic cell count should be considered when making management decisions for cows infected with S. agalactiae.

Stability of penethamate, a benzylpenicillin ester prodrug, in oily vehicles.

Penethamate (PNT) is an ester prodrug of benzylpenicillin which is marketed as dry powder for reconstitution with aqueous vehicle prior to injection. The purpose of this paper was to investigate the chemical stability of PNT in oily formulations to provide a basis for a ready-to-use (RTU) oil-based PNT formulation. The chemical stability of PNT solutions and suspensions in light liquid paraffin (LP), medium chain triglyceride (MIG), ethyl oleate (EO) and sunflower oil (SO) was investigated at 30 °C. Solid state stability of PNT powder and stability of PNT in EO suspensions with different moisture contents were also evaluated. The solubility of PNT in the oils was in order SO > EO > MIG > LP. Degradation of PNT was rapid in oily solutions and less than 10% remained after 7-15 days. Stability of PNT decreased with increase in moisture content in ethyl oleate suspensions. PNT was stable over four weeks in the solid state. Hydrolysis, due to moisture in the oil formulation is not the only degradation mechanism. PNT stability (% drug remaining) in oily suspensions after 3.5 months was in the order LP (96.2%) > MIG (95.4%) > EO (94.1%) > SO (86%). A shelf-life of up to 5.5 years at 30 °C may be achieved for PNT suspension in these oils.

Short communication: effects of systemic treatment with penethamate hydriodide on udder health and milk yields in dry primiparous Mediterranean buffaloes (Bubalus bubalis).

The effects of penethamate hydriodide (Mamyzin, Boehringer Ingelheim, Ingelheim, Germany) on udder health and milk yields were evaluated in primiparous Mediterranean buffaloes (Bubalus bubalis). An intramuscular administration of 10 million international units was performed in 20 buffaloes at 7 d precalving (treatment group; TG), and 20 animals were enrolled as the control group (CG). Evening milk samplings were performed at 10, 30, and 60 d in milk (DIM). Somatic cell count (SCC) values were evaluated on composite milk samples, whereas bacteriological culture and California Mastitis Test were performed on quarter milk. Daily milk yields were recorded after all milkings. After 60 DIM, composite milk samples from each animal were collected for monthly SCC and bacteriological culture until drying off. Statistically significant differences were found between the prevalence of mastitic quarters in the 2 groups at 10 and 30 DIM, and between the incidence of mastitic animals during the examined period (TG: 4/20, 20% vs. CG: 10/20, 50%). Even though lower and higher values of SCC and milk yields were found in TG during each sampling, statistically significant differences were only found at 30 (SCC) and 60 DIM (milk yields). In our study, the antibiotic administration precalving showed good bactericidal activity against the most common udder-specific pathogens that cause mastitis in primiparous Mediterranean buffaloes, and greater efficacy was observed at 10 and 30 DIM compared with 60 DIM. Given the significant decrease in SCC and increase in yields achieved, use of this antibiotic could be economically beneficial in buffalo breeding.

Pathogen group-specific risk factors for intramammary infection in treated and untreated dairy heifers participating in a prepartum antimicrobial treatment trial.

Heifer mastitis is a well-known problem, with several pathogens being involved. Several generic risk factors associated with the likelihood of intramammary infections (IMI) in fresh dairy heifers have been identified before. Yet, a need exists to identify pathogen group-specific factors, as the effect of (groups of) pathogens on udder health and milk yield is different. The aim of the present study was to identify pathogen group-specific risk factors for IMI in heifers participating in a prepartum antimicrobial treatment trial, allowing us to test the hypothesis that different factors are of importance between treated and untreated control heifers as well. Data from a clinical trial in which end-term heifers were treated systemically (over 3 consecutive days) 2 wk before calving with penethamate hydriodide (n=76) or remained untreated (n=73), were available. Several potential risk factors at the herd, heifer, and quarter level were recorded in the first 3 d in milk. Quarters from untreated heifers supplemented with ≥4 mg of selenium/d prepartum were significantly less likely to be infected with coagulase-negative staphylococci (CNS), whereas quarters were more likely to be infected with CNS when assistance during calving was needed. Udder edema before calving significantly decreased the odds of IMI with major pathogens. In treated heifers, no factors were detected that were associated with the likelihood of CNS IMI, whereas quarters from heifers were significantly more likely to be infected with major pathogens when they were housed in the calving pen more than 1 d and when they had been in contact with the lactating cows before calving. The risk factors for IMI that were identified in treated heifers were different than those in untreated heifers, independent of the pathogen group that was considered. It looks as if prepartum treatment not only changed the likelihood of infection, but also the factors that were associated with infection. However, except for treated heifers with an IMI with major pathogens, only a small proportion of the variation could be explained in the final models. Therefore, factors other than those that were studied could explain the likelihood of infection.

Prospective, multicenter clinical trial to validate new products for skin tests in the diagnosis of allergy to penicillin.

BACKGROUND: Allergy to penicillin is the most commonly reported type of drug hypersensitivity. Diagnosis is currently confirmed using skin tests with benzylpenicillin reagents, ie, penicilloyl-polylysine (PPL) as the major determinant of benzylpenicillin and benzylpenicillin, benzylpenicilloate and benzylpenilloate as a minor determinant mixture (MDM). OBJECTIVE: To synthesize and assess the diagnostic capacity of 2 new benzylpenicillin reagents in patients with immediate hypersensitivity reactions to B-lactams: benzylpenicilloyl octa-L-lysine (BP-OL) as the major determinant and benzylpenilloate (penilloate) as the minor determinant. METHODS: Prospective multicenter clinical trial performed in 18 Spanish centers. Efficacy was assessed by detection of positive skin test results in an allergic population and negative skin test results in a nonallergic, drug-exposed population. Sensitivity, specificity, and negative and positive predictive values were determined. RESULTS: The study sample comprised 94 allergic patients: 31 (35.23%) presented anaphylaxis, 4 (4.55%) anaphylactic shock, 51 (58.04%) urticaria, and 2 (2.27%) no specific condition. The culprit 8-lactams were amoxicillin in 63 cases (71.60%), benzypencillin in 14 cases (15.89%), cephalosporins in 2 cases (2.27%), other drugs in 3 cases (3.42%), and unidentified agents in 6 cases (6.82%). The results of testing with BP-OL were positive in 46 cases (52.3%); the results of testing with penilloate were positive in 33 cases (37.5%). When both reagents were taken into consideration, sensitivity reached 61.36% and specificity 100%. Skin testing with penilloate was significantly more often negative when the interval between the reaction and the study was longer. CONCLUSIONS: The sensitivity of BP-OL and penilloate was 61%. Considering that amoxicillin was the culprit drug in 71% of reactions, these results indicate that most patients were allergic to the whole group of penicillins. These data support the use of benzylpenicillin determinants in the diagnosis of allergy to beta-lactams, even in predominantly amoxicillin-allergic populations.

Systemic prepartum treatment of end-term dairy heifers with penethamate hydriodide: effect on udder health, milk yield, and culling until 120 days in milk.

Prepartum intramammary treatment with antimicrobials of end-term dairy heifers has frequently been proposed as a practice to reduce the prevalence of intramammary infections (IMI) at calving. From a safety standpoint for both animal and administrator, systemic treatment is preferred. A clinical trial was conducted on heifers from 10 well-managed, commercial dairy farms with a low prevalence of heifer mastitis. The aim was to assess both the short- and long-term effects of a systemic prepartum therapy with penethamate hydriodide on udder health and milk production. Because it was hypothesized that some herds would benefit more from this treatment than others, specific herd-level information was collected before the start of the actual trial to screen for and explain potential herd-specific treatment effects. Further, the effect of treatment on antimicrobial susceptibility of staphylococcal isolates was monitored. End-term heifers were either treated systemically (over 3 consecutive days) 2 wk before expected calving date with penethamate hydriodide (n=76) or remained untreated (n=73). Systemic prepartum treatment of end-term heifers with penethamate hydriodide resulted in fewer IMI in early lactation. However, all 6 cases of clinical mastitis in early lactation occurred in the treatment group [Streptococcus uberis (n=1), Corynebacterium bovis (n=1), Staphylococcus aureus (n=1); 1 sample was contaminated; 2 samples remained culture negative]. No long-term treatment effects (from 4 to 120 d in milk) on milk production, udder health, or culling hazard during later lactation were detected, although treated heifers belonging to herds classified as having low-yielding heifers out-produced the control heifers. Moreover, penicillin susceptibility of staphylococci isolated from milk samples of treated or control heifers did not differ. Herds with a low prevalence of heifer mastitis are not likely to benefit from prepartum systemic antimicrobial treatment of the end-term heifers.

Characterization of ß-lactamase enzyme activity in bacterial lysates using MALDI-mass spectrometry.

Plasmid-encoded ß-lactamases are a major reason for antibiotic resistance in gram negative bacteria. These enzymes hydrolyze the ß-lactam ring structure of certain ß-lactam antibiotics, consequently leading to their inactivation. The clinical situation demands for specific first-line antibiotic therapy combined with a quick identification of bacterial strains and their antimicrobial susceptibility. Strategies for the identification of ß-lactamase activity are often cumbersome and usually lack sensitivity and specificity. The current work demonstrates that matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) is an ideal tool for these analytical investigations. Herein, we describe a fast and specific assay to determine ß-lactamase activity in bacterial lysates. The feasibility of the analytical read-out was demonstrated on a MALDI-triple quadrupole (QqQ) and a MALDI time-of-flight (TOF) instrument, and the results allow the comparison of both approaches. The assay specifically measures enzyme-mediated, time-dependent hydrolysis of the ß-lactam ring structure of penicillin G and ampicillin and inhibition of hydrolysis by clavulanic acid for clavulanic acid susceptible ß-lactamases. The assay is reproducible and builds the basis for future in-depth investigations of ß-lactamase activity in various bacterial strains by mass spectrometry.

Diagnosis of immediate hypersensitivity to ß-lactam antibiotics can be made safely with current approaches.

BACKGROUND: Diagnosing immediate hypersensitivity to ß-lactam antibiotics is still a significant problem. Recently, a new penicillin testing reagent was introduced to the market. In this study, the recommendations of the European Network of Drug Allergy (ENDA) for the diagnosis of immediate reactions to ß-lactams were followed, and the negative predictive value of this approach with currently available reagents was assessed. METHODS: Eighty patients (age range: 6-74 years) with a history of immediate reactions to ß-lactams were included. All cases underwent skin testing with benzylpenicilloyl-polylysine (PPL) and minor determinant mixture (MDM), followed by the culprit drug if necessary. If this step was negative, a drug provocation test was offered. If this step also yielded a negative result, then the patients were recommended to use ß-lactam antibiotics in future whenever their use was indicated. RESULTS: Overall, 29 patients (36.2%) were diagnosed as ß-lactam allergic. The majority of the cases (72.4%) were diagnosed by positive skin tests to either PPL or MDM, whereas 10.3% were diagnosed by skin testing with culprit drugs and 17.2% with drug provocation tests. Regarding the use of the tested drug in the long term, almost half of the contacted patients had had an indication to use the tested drug and the majority had taken the whole course without problems. CONCLUSIONS: Although currently available new penicillin tests provide sufficient allergy data, all the steps recommended by ENDA should be followed in the diagnosis of immediate reactions to ß-lactams. If these steps are negative, the patients usually tolerate ß-lactams and only a few develop mild, non-life-threatening reactions in the long term.

Alkali-treated penicillin G solution is a better option than penicillin G as an alternative source of minor determinants for penicillin skin test.

Both benzylpenicilloyl-polylysine (PPL) and minor determinant mixture (MDM) are the recommended standard reagents for penicillin skin testing. However, penicillin G is commonly suggested as an alternative source of minor determinants. This study evaluated the accuracy of penicillin G and alkali-treated penicillin G compared with the standardized MDM for skin testing. Sixty-eight patients with histories of allergies to penicillin or semisynthetic penicillins were skin tested with commercial Kit penicillin allergenic determinants (DAP) (PPL and DAP-MDM; Diater Laboratorios, Madrid, Spain). The in-house MDM (IH-MDM), prepared by alkali-treated aged penicillin, and fresh penicillin G sodium (PGs) were tested alongside DAP-MDM. Positive penicillin skin test results were identified in 22 patients (32.4%) using commercial reagents (PPL+ DAP-MDM) and 19 of them reacted to DAP-MDM alone or together with PPL. The accuracy of IH-MDM and PGs compared with DAP-MDM was 89.7 and 76.5%, respectively. Our study shows that alkali-treated penicillin G is a better option than penicillin G as an alternative source of MDM for skin testing in case the commercialized MDM is not available. Minor determinants play a significant role for penicillin allergy in Thailand and should be included in the penicillin skin test panel to verify suspected cases of penicillin allergy. (ClinicalTrials.gov number: NCT00789217).

Pre-formulation and chemical stability studies of penethamate, a benzylpenicillin ester prodrug, in aqueous vehicles.

Penethamate (PNT) is a diethylaminoethyl ester prodrug of benzylpenicillin used to treat bovine mastitis via the intramuscular route. Because of its instability, PNT products must be reconstituted before administration and the reconstituted injection has a short shelf life (7 days at 2-8°C). The purpose of this paper was to investigate whether the stability of PNT can be improved in order to achieve a chemically stable ready-to-use aqueous-based PNT formulation or at least to extend the shelf life of the reconstituted suspension. A chemical stability study of PNT in aqueous-based solutions as a function of pH, buffer strength, solvent mixtures and temperature, supported by studies of its solubility in mixed solvents, allowed predictions of the shelf life of PNT solution and suspension formulations. PNT degraded in aqueous solutions by several pathways over the pH range 2.0-9.3 with a V-shaped pH-rate profile and a minimum pH of around 4.5. The stability of PNT solutions in mixed solvents was greater than in aqueous solutions. For example, in propylene glycol:citrate buffer (60:40, v/v, pH 4.5), the half-life of PNT was 4.3 days compared with 1.8 days in aqueous buffer. However, solubility of PNT in the mixed solvent was higher than that in aqueous solution and this had an adverse effect on the stability of suspensions. By judicious choosing of pH and mixed solvent, it is possible to achieve a storage life of a PNT suspension of 5.5 months at 5°C, not sufficient for a ready-to-use product but a dramatic improvement in the storage life of the reconstituted product.

Direct evidence for the formation of diastereoisomeric benzylpenicilloyl haptens from benzylpenicillin and benzylpenicillenic acid in patients.

Covalent binding to proteins to form neoantigens is thought to be central to the pathogenesis of penicillin hypersensitivity reactions. We have undertaken detailed mass spectrometric studies to define the mechanism and protein chemistry of hapten formation from benzylpenicillin (BP) and its rearrangement product, benzylpenicillenic acid (PA). Mass spectrometric analysis of human serum albumin exposed to BP and PA in vitro revealed that at low concentrations (drug protein molar ratio 0.001:1) and during short time incubations BP and PA selectively target different residues, Lys199 and Lys525, respectively. Molecular modeling showed that the selectivity was a function of noncovalent interaction before covalent modification. With increased exposure to higher concentrations of BP and PA, multiple epitopes were detected on albumin, demonstrating that the multiplicity of hapten formation is a function of time and concentration. More importantly, we have demonstrated direct evidence that PA is a hapten accounting for the diastereoisomeric BP antigen formation in albumin isolated from the blood of patients receiving penicillin. Furthermore, PA was found to be more potent than BP with respect to stimulation of T cells from patients with penicillin hypersensitivity, illustrating the functional relevance of diastereoisomeric hapten formation.

Non-antibiotic properties of tetracyclines as anti-allergy and asthma drugs.

All available therapies for human allergic disease target IgE mediated pathologic responses after IgE has been produced. We are developing tetracyclines as anti-allergy drugs to prevent IgE production, based on our findings that minocycline or doxycycline treatment of allergic asthmatic humans significantly improves their asthma symptoms, reduces their oral steroid requirements, and strongly suppresses their ongoing IgE responses (ELISA, mast cell mediated cutaneous late phase responses); the tetracyclines also strongly suppress peak IgE responses of BPO-KLH sensitized mice (ELISPOT assay, ELISA, skin tests). The antibiotic activity of the tetracyclines is not required for suppression of IgE responses; inclusion of minocycline or doxycycline in sterile culture prevents anti-CD40/IL-4 mediated induction of memory IgE responses by PBMC of allergic asthmatic patients (ELISA), and induction of specific memory IgE responses by spleen cells of BPO-KLH sensitized mice (ELISPOT assay, ELISA). The tetracyclines affect an epsilon specific pathway because IgM, IgG and IgA responses did not decrease. Further, in humans, DTH responses to recall antigens did not decrease. In related studies, we found that two distinct T cell subsets: CD4+CD60 negative and CD8+CD60+ (CD60 is a ganglioside) (humans) and CD4+ Asialo GM1 ganglioside negative and CD8+Asialo GM1 ganglioside+ (mice), both are required for induction of memory IgE responses. Phosphorylated (phos) p38 MAP kinase, but not phos ERK or phos JNK expression by CD4+ and CD8+, including CD8+CD60+, T cells is increased in allergic asthmatic humans, as is IL-4 and IL-10 production. The tetracyclines appear to target T cell pathways to induce suppression of IgE responses because they suppress phos p38 MAP kinase expression by both CD4+ and CD8+, including CD8+CD60+, T cell subsets, and IL-4 and IL-10, while upregulating IL-2 and IFN gamma, and suppressing IgE responses. Our finding that tetracyclines do not require antibiotic activity to suppress IgE responses opens the door to development of new tetracycline-based and other therapeutics for human allergic disease.

Saturation mutagenesis of Acremonium chrysogenum deacetoxy/deacetylcephalosporin C synthase R308 site confirms its role in controlling substrate specificity.

Deacetoxy/deacetylcephalosporin C synthase (acDAOC/DACS) from Acremonium chrysogenum is a bifunctional enzyme that catalyzes both the ring-expansion of penicillin N to deacetoxycephalosporin C and the hydroxylation of the latter to deacetylcephalosporin C. The R308 residue located in close proximity to the C-terminus of acDAOC/DACS was mutated to the other 19 amino acids. In the resulting mutant pool, R308L, R308I, R308T and R308V showed significant improvement in their ability to convert penicillin analogs, thus confirming the role of R308 in controlling substrate selectivity, the four amino acids all possess short aliphatic sidechains that may improve hydrophobic interactions with the substrates. The mutant R308I showed the highest reactivity for penicillin G, with 3-fold increase in k(cat)/K(m) ratio and 7-fold increase in relative activity.

Regulatory functions of hapten-reactive helper and suppressor T lymphocytes. II. Selective inactivation of hapten-reactive suppressor T cells by hapten-nonimmunogenic copolymers of D-amino acids, and its application to the study of suppressor T-cell effect on helper T-cell development.

An experimental condition was established in vivo for selectively eliminating hapten-reactive suppressor T-cell activity generated in mice primed with a para-azobenzoate (PAB)-mouse gamma globulin (MGG)-conjugate and treated with PAB-nonimmunogenic copolymer of D-amino acids (D- glutamic acid and D-lysine; D-GL). The elimination of suppressor T-cell activity with PAB-D-GL treatment from the mixed populations of hapten- reactive suppressor and helper T cells substantially increased apparent helper T-cell activity. Moreover, the inhibition of PAB-reactive suppressor T-cell generation by the pretreatment with PAB-D-GL before the PAB-MGG-priming increased the development of PAB-reactive helper T-cell activity. The analysis of hapten-specificity of helper T cells revealed that the reactivity of helper cells developed in the absence of suppressor T cells was more specific for primed PAB-determinants and their cross-reactivities to structurally related determinants such as meta-azobenzoate (MAB) significantly decreased, as compared with the helper T-cell population developed in the presence of suppressor T lymphocytes. In addition, those helper T cells generated in the absence of suppressor T cells were highly susceptible to tolerogenesis by PAB-D- GL. Similarly, the elimination of suppressor T lymphocytes also enhanced helper T-cell activity in a polyclonal fashion in the T-T cell interactions between benzylpenicilloyl (BPO)-reactive T cells and PAB- reactive T cells after immunization of mice with BPO-MGG-PAB. Thus inhibition of BPO-reactive suppressor T-cell development by the BPO-v-GL- pretreatment resulted in augmented generation of PAB-reactive helper T cells with higher susceptibility of tolerogenesis to PAB-D-GL. Thus, these results support the notion that suppressor T cells eventually suppress helper T-cell activity and indicate that the function of suppressor T cells related to helper T-cell development is to inhibit the increase in the specificity and apparent affinity of helper T cells in the primary immune response. The hapten-reactive suppressor and helper T lymphocytes are considered as a model system of T cells that regulate the immune response, and the potential applicability of this system to manipulating various T cell-mediated immune responses is discussed in this context.

Penicillin allergy: value of including amoxicillin as a determinant in penicillin skin testing.

BACKGROUND: Allergy to penicillins remains an important issue. Penicillin skin testing (PST) with major and minor determinants has been shown to be a highly valuable tool for identifying IgE-mediated penicillin allergy. The value of additional testing with side-chain-specific moieties from semisynthetic penicillins such as amoxicillin is not well-established in spite of the widespread use of these medications. METHODS: A retrospective review of all consecutive inpatient PST results from 1995 to 2007 comprising 1,068 patients was performed in our institution on individuals with a self-reported history of beta-lactam allergy to assess the importance of including the amoxicillin determinant in a previously validated PST panel. Descriptive statistics were performed. The PST panel included penicilloyl-polylysine, penicillin G, penicilloate, penilloate and amoxicillin. RESULTS: Of 1,068 patients, 243 (23%) had a positive skin test reaction on the PST panel. Testing with amoxicillin was positive in 30.9% of patients, the majority of whom (81%) were also positive to 1 or more standard penicillin reagents. Fourteen of the 243 positive patients (5.8%) had a positive skin test reaction only to amoxicillin. Additionally, the use of penicilloate and penilloate minor determinants in combination with penicillin G identified a greater percentage of penicillin-allergic individuals compared to using only penicillin G (22.6 vs. 6.6%), demonstrating their importance in the PST panel. CONCLUSIONS: These data indicate that the inclusion of the amoxicillin determinant appears to identify a small but important group of allergic individuals who may otherwise test negative on a PST panel.

Concentration of penicillin G in mammary tissue and secretion of end-term dairy heifers following systemic prepartum administration of penethamate hydriodide.

The aim of this study was to assess the concentration of penicillin G in mammary tissue and secretion of dry heifers following systemic administration of penethamate hydriodide. Six dairy heifers in late gestation received a single intramuscular injection of 10 g penethamate hydriodide and were sacrificed 24, 48 or 144 h after treatment. Penicillin G concentrations were measured in mammary tissue and secretion samples using HPLC. Penicillin G was detected in the udder of two animals euthanized at 24 h (mammary tissue and secretion) and at 48 h post treatment (mammary secretion only) after administration at concentrations still close to or above MIC90 values reported for the pathogens associated with heifer mastitis. Antibiotic concentration shortly after administration will have been substantially higher indicating a potential for systemic treatment with penethamate hydriodide to control prepartum intramammary infections in heifers without the disadvantages of local therapy such as teat contamination or risk of trauma for the administrator.