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Sample records for lactam bridge-cyclized alpha-melanocyte

  1. alpha-Melanocyte stimulating hormone: immunohistochemical identification and mapping in neurons of rat brain.

    PubMed Central

    Jacobowitz, D M; O'Donohue, T L

    1978-01-01

    alpha-Melanocyte stimulating hormone (alpha-melanotropin) immunofluorescence was observed in rat brain by means of a highly specific and well-characterized antibody. The hormone was contained in arcuate nucleus cell bodies and in varicose fibers. Dense populations of hormone-containing fibers were present in the septum, the nucleus interstitialis stria terminalis, and the medial preoptic, anterior hypothalamic, dorsomedial, and periventricular nuclei. Moderate numbers of fibers were seen in the paraventricular and arcuate nuclei, the amygdala, the region of the tractus diagonalis, the mammillary body, the central gray, the cuneiform nucleus, and the nucleus of the solitary tract. There is an interesting correlation of alpha-melanocyte stimulating hormone fibers with regions of noradrenergic axonal projections and terminal fields. Images PMID:366617

  2. Mapping of alpha-melanocyte-stimulating hormone-like immunoreactivity in the cat brainstem.

    PubMed

    Coveñas, R; de León, M; Narváez, J A; Aguirre, J A; Tramu, G

    2000-04-01

    The distribution of alpha-melanocyte-stimulating hormone-like immunoreactive structures was studied in the brainstem of the cat using an indirect immunoperoxidase technique. Immunoreactivity was observed in several brainstem nuclei of the cat in which no immunoreactivity had been previously reported. Immunoreactive fibres were observed in the following; the inferior central nucleus; the pontine gray nuclei; the Kölliker-Fuse nucleus; the motor trigeminal nucleus, the anteroventral cochlear nucleus; the abducens nucleus; the retrofacial nucleus; the superior, lateral, inferior, and medial vestibular nuclei; the lateral nucleus of the superior olive; the external cuneate nucleus; the nucleus of the trapezoid body; the postpyramidal nucleus of the raphe; the medial accessory inferior olive; the dorsal accessory nucleus of the inferior olive; the nucleus ambiguus; the principal nucleus of the inferior olive; the preolivary nucleus; the nucleus ruber; the substantia nigra; and in the area postrema. Our results point to a more widespread distribution of alpha-melanocyte-stimulating hormone-like immunoreactive structures in the cat brainstem than that reported in previous studies carried out in the same region of the cat, rat and humans.

  3. Imaging human melanoma using a novel Tc-99m-labeled lactam bridge-cyclized alpha-MSH peptide.

    PubMed

    Liu, Liqin; Xu, Jingli; Yang, Jianquan; Feng, Changjian; Miao, Yubin

    2016-10-01

    In this study, the human melanoma targeting property of (99m)Tc(EDDA)-HYNIC-AocNle-CycMSHhex {hydrazinonicotinamide-8-aminooctanoic acid-Nle-c[Asp-His-DPhe-Arg-Trp-Lys]-CONH2} was determined in M21 human melanoma-xenografts to demonstrate its potential for human melanoma imaging. The IC50 value of HYNIC-AocNle-CycMSHhex was 0.48±0.01nM in M21 human melanoma cells (1281receptors/cell). The M21 human melanoma uptake of (99m)Tc(EDDA)-HYNIC-AocNle-CycMSHhex was 4.03±1.25, 3.26±1.23 and 3.36±1.48%ID/g at 0.5, 2 and 4h post-injection, respectively. Approximately 92% of injected dose cleared out the body via urinary system at 2h post-injection. (99m)Tc(EDDA)-HYNIC-AocNle-CycMSHhex showed high tumor/blood, tumor/muscle and tumor/skin uptake ratios after 2h post-injection. The M21 human melanoma-xenografted tumor lesions were clearly visualized by SPECT/CT using (99m)Tc(EDDA)-HYNIC-AocNle-CycMSHhex as an imaging probe at 2h post-injection. Overall, (99m)Tc(EDDA)-HYNIC-AocNle-CycMSHhex exhibited favorable human melanoma imaging property, highlighting its potential as an imaging probe for human metastatic melanoma detection. PMID:27568083

  4. Alpha-Melanocyte Stimulating Hormone: An Emerging Anti-Inflammatory Antimicrobial Peptide

    PubMed Central

    Singh, Madhuri; Mukhopadhyay, Kasturi

    2014-01-01

    The alpha-melanocyte stimulating hormone (α-MSH) is a neuropeptide belonging to the melanocortin family. It is well known for its anti-inflammatory and antipyretic effects and shares several characteristics with antimicrobial peptides (AMPs). There have been some recent reports about the direct antimicrobial activity of α-MSH against various microbes belonging to both fungal and bacterial pathogens. Similar to α-MSH's anti-inflammatory properties, its C-terminal residues also exhibit antimicrobial activity parallel to that of the entire peptide. This review is focused on the current findings regarding the direct antimicrobial potential and immunomodulatory mechanism of α-MSH and its C-terminal fragments, with particular emphasis on the prospects of α-MSH based peptides as a strong anti-infective agent. PMID:25140322

  5. GABAergic agents prevent alpha-melanocyte stimulating hormone induced anxiety and anorexia in rats.

    PubMed

    Rao, T Lakshmi; Kokare, Dadasaheb M; Sarkar, Sumit; Khisti, Rahul T; Chopde, Chandrabhan T; Subhedar, Nishikant

    2003-12-01

    Alpha-melanocyte stimulating hormone (alpha-MSH) is a hypothalamic peptide believed to play a tonic inhibitory role in feeding and energy homeostasis. Systemic administration of alpha-MSH is known to produce anorexia and anxiety. Since synaptic contacts between gamma-aminobutyric acid (GABA)ergic terminals and alpha-MSH neurons in the hypothalamus have been reported, the present work was undertaken to refine our knowledge on the role of GABAergic systems in anxiety and anorexia induced by intracerebroventricular (icv) administration of alpha-MSH in rats. The anxiety was assessed by elevated plus maze, and spontaneous food consumption was monitored during dark cycle. Prior administration of diazepam and muscimol that promote the function of GABA(A) receptors reversed the anxiogenic response and decreased food intake elicited by alpha-MSH. In contrast, bicuculline, the GABA(A) receptor antagonist, not only enhanced the effects of alpha-MSH but also prevented the influence of GABAergic drugs on alpha-MSH-induced anorexia and anxiety. These findings suggest that alpha-MSH-induced anxiety and anorexia are due to its negative influence on GABAergic system.

  6. Alpha-melanocyte-stimulating hormone down-regulates CXC receptors through activation of neutrophil elastase.

    PubMed

    Manna, Sunil K; Sarkar, Abira; Sreenivasan, Yashin

    2006-03-01

    Considering the role of interleukin-8 (IL-8) in a large number of acute and chronic inflammatory diseases, the regulation of IL-8-mediated biological responses is important. Alpha-melanocyte-stimulating hormone (alpha-MSH), a tridecapeptide, inhibits most forms of inflammation by an unknown mechanism. In the present study, we have found that alpha-MSH interacts predominantly with melanocortin-1 receptors and inhibits several IL-8-induced biological responses in macrophages and neutrophils. It down-regulated receptors for IL-8 but not for TNF, IL-4, IL-13 or TNF-related apoptosis-inducing ligand (TRAIL) in neutrophils. It down-regulated CXCR type 1 and 2 but not mRNA levels. alpha-MSH did not inhibit IL-8 binding in purified cell membrane or affinity-purified CXCR. IL-8 or anti-CXCR Ab protected against alpha-MSH-mediated inhibition of IL-8 binding. The level of neutrophil elastase, a specific serine protease, but not cathepsin G or proteinase 3 increased in alpha-MSH-treated cells, and restoration of CXCR by specific neutrophil elastase or serine protease inhibitors indicates the involvement of elastase in alpha-MSH-induced down-regulation of CXCR. These studies suggest that alpha-MSH inhibits IL-8-mediated biological responses by down-regulating CXCR through induction of serine protease and that alpha-MSH acts as a potent immunomodulator in neutrophil-driven inflammatory distress. PMID:16479540

  7. Oral Bifidobacterium longum expressing alpha-melanocyte-stimulating hormone to fight experimental colitis.

    PubMed

    Wei, Pijin; Yang, Yan; Liu, Zhaobing; Huang, Junli; Gong, Yahui; Sun, Hanxiao

    2016-07-01

    The oral delivery of peptides is a highly attractive treatment approach. However, the harsh environment of the gastrointestinal tract limits its application. Here, we utilize Bifidobacterium as a delivery system to orally deliver a potent anti-inflammatory but short duration peptide alpha-melanocyte-stimulating hormone (α-MSH) against experimental colitis. The aim of our study was to facilitate the efficient oral delivery of α-MSH. We designed a vector of pBDMSH and used it to construct a Bifidobacterium longum expressing α-MSH. We then determined the bioactivity of recombinant Bifidobacterium in lipopolysaccharide-induced inflammatory models of HT-29 cells. Finally, we used Bifidobacterium expressing α-MSH against dextran sulfate sodium (DSS)-induced ulcerative colitis mice. Results based on the myeloperoxidase activity, the levels of inflammatory cytokines TNF-α, IL-1β, IL-6, and IL-10 and the histological injury of colon tissue reveal recombinant Bifidobacterium was efficient in attenuating DSS-induced ulcerative colitis, suggesting an alternative way to use Bifidobacterium as a delivery system to deliver α-MSH for DSS-induced ulcerative colitis therapy. PMID:26673899

  8. Alpha-melanocyte-stimulating hormone reduces endotoxin-induced liver inflammation.

    PubMed

    Chiao, H; Foster, S; Thomas, R; Lipton, J; Star, R A

    1996-05-01

    Alpha-Melanocyte-stimulating hormone (MSH) is a potent anti-inflammatory agent in many models of inflammation, suggesting that it inhibits a critical step common to different forms of inflammation. We showed previously that alpha-MSH inhibits nitric oxide (NO) production in cultured macro-phages. To determine how alpha-MSH acts in vivo, we induced acute hepatic inflammation by administering endotoxin (LPS) to mice pretreated with Corynebacterium parvum, alpha-MSH prevented liver inflammation even when given 30 min after LPS administration. To determine the mechanisms of action of alpha-MSH, we tested its influence on NO, infiltrating inflammatory cells, cytokines, and chemokines. Alpha-MSH inhibited systemic NO production, hepatic neutrophil infiltration, and increased hepatic mRNA abundance for TNF alpha, and the neutrophil and monocyte chemokines (KC/IL-8 and MCP-1). We conclude that alpha-MSH prevents LPS-induced hepatic inflammation by inhibiting production of chemoattractant chemokines which then modulate infiltration of inflammatory cells. Thus, alpha-MSH has an effect very early in the inflammatory cascade. PMID:8621792

  9. Nocturnal plasma melatonin and alpha-melanocyte stimulating hormone levels during exacerbation of multiple sclerosis.

    PubMed

    Sandyk, R; Awerbuch, G I

    1992-01-01

    The pineal gland has been implicated recently in the pathogenesis of multiple sclerosis (MS). To investigate this hypothesis further, we studied nocturnal plasma melatonin levels and the presence or absence of pineal calcification (PC) on CT scan in a cohort of 25 patients (5 men, 20 women; mean age: 41.1 years; SD = 11.1; range: 27-72) who were admitted to a hospital Neurology service for exacerbation of symptoms. Plasma alpha-melanocyte stimulating hormone (alpha-MSH) estimations were included in the study since there is evidence for a feedback inhibition between alpha-MSH and melatonin secretion. Abnormal melatonin levels were found in 13 patients (52.0%), 11 of whom had nocturnal levels which were below the daytime values (i.e., < 25 pg/ml). Although melatonin levels were unrelated to the patient's age and sex, there was a positive correlation with age of onset of symptoms (p < .0001) and an inverse correlation with the duration of illness (p < .05). PC was noted in 24 of 25 patients (96%) underscoring the pathogenetic relationship between MS and the pineal gland. Alpha-MSH levels were undetectable in 15 patients (60.0%), low in two patients (8.0%), normal in seven patients (28.0%), and elevated in one patient (4.0%). Collectively, abnormal alpha-MSH levels were found in over 70% of patients. These findings support the hypothesis that MS may be associated with pineal failure and suggest, furthermore, that alterations in the secretion of alpha-MSH also occur during exacerbation of symptoms. The relevance of these findings to the pathogenesis of MS are discussed. PMID:1305632

  10. DOTA alpha-melanocyte-stimulating hormone analogues for imaging metastatic melanoma lesions.

    PubMed

    Froidevaux, Sylvie; Calame-Christe, Martine; Sumanovski, Lazar; Tanner, Heidi; Eberle, Alex N

    2003-06-01

    Scintigraphic imaging of metastatic melanoma lesions requires highly tumor-specific radiopharmaceuticals. Because both melanotic and amelanotic melanomas overexpress melanocortin-1 receptors (MC1R), radiolabeled analogues of alpha-melanocyte-stimulating hormone (alpha-MSH) are potential candidates for melanoma diagnosis. Here, we report the in vivo performance of a newly designed octapeptide analogue, [betaAla(3), Nle(4), Asp(5), D-Phe(7), Lys(10)]-alpha-MSH(3-10) (MSH(OCT)), which was conjugated through its N-terminal amino group to the metal chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) to enable incorporation of radiometals (e.g., indium-111) into the peptide. DOTA-MSH(OCT) displayed high in vitro MC1R affinity (IC(50) 9.21 nM). In vivo [(111)In]DOTA-MSH(OCT) exhibited a favorable biodistribution profile after injection in B16-F1 tumorbearing mice. The radiopeptide was rapidly cleared from blood through the kidneys and, most importantly, accumulated preferentially in the melanoma lesions. Lung and liver melanoma metastases could be clearly imaged on tissue section autoradiographs 4 h after injection of [(111)In]DOTA-MSH(OCT). A comparative study of [(111)In]DOTA-MSH(OCT) with [(111)In]DOTA-[Nle(4), D-Phe(7)]-alpha-MSH ([(111)In]-DOTA-NDP-MSH) demonstrated the superiority of the DOTA-MSH(OCT) peptide, particularly for the amount of radioactivity taken up by nonmalignant organs, including bone, the most radiosensitive tissue. These results demonstrate that [(111)In]DOTA-MSH(OCT) is a promising melanoma imaging agent.

  11. Alpha-melanocyte stimulating hormone ameliorates disease activity in an induced murine lupus-like model.

    PubMed

    Botte, D A C; Noronha, I L; Malheiros, D M A C; Peixoto, T V; de Mello, S B V

    2014-08-01

    Alpha-melanocyte stimulating hormone (α-MSH) is a neuropeptide exhibiting anti-inflammatory activity in experimental models of autoimmune diseases. However, no studies thus far have examined the effects of α-MSH on systemic lupus erythematosus (SLE). This study aimed to determine the effects of an α-MSH agonist in induced murine lupus. Here we employed female Balb/cAn mice in which lupus was induced by pristane. Groups of lupus animals were treated daily with the α-MSH analogue [Nle4, DPhe7]-α-MSH (NDP-MSH) (1·25 mg/kg) injected intraperitoneally or saline for 180 days. Normal animals comprised the control group. Arthritis incidence, plasma immunoglobulin (Ig)G isotypes, anti-nuclear antibodies (ANA) and plasma cytokines were evaluated. Renal function was assessed by proteinuria and histopathological lesion. Glomerular levels of IgG, α-smooth muscle actin (α-SMA), inducible nitric oxide synthase (iNOS), C3, CD3, melanocortin receptors (MCR)1, corticotrophin-releasing factor (CRF) and α-MSH was estimated by immunohistochemistry. When compared with normal controls, lupus animals exhibited increased arthritis, IgG levels, ANA, interleukin (IL)-6, IL-10, proteinuria and mesangial cell proliferation together with glomerular expression of α-SMA and iNOS. Glomerular expression of MCR1 was reduced in lupus animals. NDP-MSH treatment reduced arthritis scores by 70% and also diminished IgG1 and IgG2a levels and ANA incidence. In the glomerulus, NDP-MSH treatment reduced cellularity by 50% together with reducing IgG deposits, and expression levels of α-SMA, iNOS and CRF were also all decreased. Taken together, our results suggest for the first time that α-MSH treatment improves several parameters of SLE disease activity in mice, and indicate that this hormone is an interesting potential future treatment option.

  12. Melanoma Therapy with Rhenium-Cyclized Alpha Melanocyte Stimulating Hormone Peptide Analogs

    SciTech Connect

    Thomas P Quinn

    2005-11-22

    Malignant melanoma is the 6th most commonly diagnosed cancer with increasing incidence in the United States. It is estimated that 54,200 cases of malignant melanoma will be newly diagnosed and 7,600 cases of death will occur in the United States in the year 2003 (1). At the present time, more than 1.3% of Americans will develop malignant melanoma during their lifetime (2). The average survival for patients with metastatic melanoma is about 6-9 months (3). Moreover, metastatic melanoma deposits are resistant to conventional chemotherapy and external beam radiation therapy (3). Systematic chemotherapy is the primary therapeutic approach to treat patients with metastatic melanoma. Dacarbazine is the only single chemotherapy agent approved by FDA for metastatic melanoma treatment (5). However, the response rate to Dacarbazine is only approximately 20% (6). Therefore, there is a great need to develop novel treatment approaches for metastatic melanoma. The global goal of this research program is the rational design, characterization and validation of melanoma imaging and therapeutic radiopharmaceuticals. Significant progress has been made in the design and characterization of metal-cyclized radiolabeled alpha-melanocyte stimulating hormone peptides. Therapy studies with {sup 188}Re-CCMSH demonstrated the therapeutic efficacy of the receptor-targeted treatment in murine and human melanoma bearing mice (previous progress report). Dosimetry calculations, based on biodistribution data, indicated that a significant dose was delivered to the tumor. However, {sup 188}Re is a very energetic beta-particle emitter. The longer-range beta-particles theoretically would be better for larger tumors. In the treatment of melanoma, the larger primary tumor is usually surgically removed leaving metastatic disease as the focus of targeted radiotherapy. Isotopes with lower beta-energies and/or shorter particle lengths should be better suited for targeting metastases. The {sup 177}Lu

  13. The Alpha-Melanocyte-Stimulating Hormone Suppresses TLR2-Mediated Functional Responses through IRAK-M in Normal Human Keratinocytes

    PubMed Central

    Ryu, Sunhyo; Johnson, Andrew; Park, Yoonkyung; Kim, Beomjoon; Norris, David; Armstrong, Cheryl A.; Song, Peter I.

    2015-01-01

    Alpha-melanocyte stimulating hormone (α-MSH) is a highly conserved 13-aa neuropeptide derived from pro-opiomelanocortin by post-translational processing, which has been reported to exhibit potent anti-inflammatory activity and a wide range of immunosuppressive activities in the skin. However, the regulatory effect of α-MSH is not completely clear in cutaneous innate immunity. In this study, we investigate the functional regulation of α-MSH in TLR2-mediated inflammatory responses in normal human keratinocytes (HKs). α-MSH pretreatment down-regulated the Staphylococcus aureus LTA-induced expression of both TLR2 and IL-8 as well as NF-κB nuclear translocation in HK cells. The inhibitory effect of α-MSH was blocked by agouti signaling protein (ASP), an α-MSH receptor-1 antagonist. To investigate the mechanism of this response in more detail, siRNA of IRAK-M, a negative regulator of TLR signaling, was utilized in these studies. The α-MSH suppressive effect on IL-8 production and NF-κB transactivation was inhibited by IRAK-M siRNA transfection in HK cells. These results indicate that α-MSH is capable of suppressing keratinocyte TLR2-mediated inflammatory responses induced by S. aureus-LTA, thus demonstrating another novel immunomodulatory activity of α-MSH in normal human keratinocytes. PMID:26309029

  14. Reducing renal uptake of 90Y- and 177Lu-labeled alpha-melanocyte stimulating hormone peptide analogues

    SciTech Connect

    Miao, Yubin; Fisher, Darrell R.; Quinn, Thomas P.

    2006-06-15

    The purpose of this study was to improve the tumor-to-kidney uptake ratios of 90Y- and 177Lu-[1,2,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid-Re-Cys,D-Phe,Arg]alpha-melanocyte stimulating hormone (DOTA-RE(Arg)CCMSH), through coupling a negatively charged glutamic acid (Glu) to the peptide sequence. A new peptide of DOTA-Re(Glu,Arg)CCMSH was designed, synthesized and labeled with 90Y and 177Lu. Pharmacokinetics of 90Y- and 177Lu-DOTA-RE(Glu,Arg)CCNSH were determined in B16/F1 murine melanoma-bearing C57 mice. Both exhibited significantly less renal uptake than 90Y- and 177Lu-DOTA-Re(Arg)CCMSH at 30 min and at 2, 3, and 24 h after dose administration. The renal uptake values of 90Y- and 177Lu-DOTA-Re(Glu,Arg)CCMSH were 28.16% and 28.81% of those of 90Y- and 177Lu-DOTA-RE(Arg)CCMSH, respectively, at 4 hr post-injection. We also showed higher tumor-to-kidney uptake ratios 2.28 and 1.69 times that of 90Y- and 177Lu-DOTA-Re(Arg)CCMSH, respectively, at 4 h post-injection. The90Y- and 177Lu-DOTA-Re(Glu,Arg)CCMSH activity accumulation was low in normal organs except for kidneys. Coupling a negatively charged amino acid (Glu) to the CCMSH peptide sequence dramatically reduced the renal uptake values and increased the tumor-to-kidney uptake ratios of 90Y- and 177Lu-DOTA-Re(Glu,Arg)CCMSH, facilitating their potential applications as radiopharmaceuticals for targeted radionuclide therapy of melanoma.

  15. 203Pb-Labeled Alpha-Melanocyte-Stimulating Hormone Peptide as an Imaging Probe for Melanoma Detection

    SciTech Connect

    Yubin, Miao; Figueroa, Said D.; Fisher, Darrell R.; Moore, Herbert A.; Testa, Richard F.; Hoffman, Timothy J.; Quinn, Thomas P.

    2008-05-01

    Abbreviations: a-MSH; alpha melanocyte stimulating hormone, DOTA; 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, Re(Arg11)CCMSH; DOTA-[Cys3,4,10, D-Phe7, Arg11]a-MSH3-13, NDP; [Nle4,d-Phe7] a-MSH3-13. Abstract Peptide-targeted alpha therapy with 200 mCi of 212Pb-DOTA-Re(Arg11)CCMSH cured 45% of B16/F1 murine melanoma-bearing C57 mice in a 120-day study, highlighting its melanoma treatment potential. However, there is a need to develop an imaging surrogate for patient specific dosimetry and to monitor the tumor response to 212Pb-DOTA-Re(Arg11)CCMSH therapy. The purpose of this study was to evaluate the potential of 203Pb-DOTA-Re(Arg11)CCMSH as a matched-pair SPECT imaging agent for 212Pb-DOTA-Re(Arg11)CCMSH. Method: DOTA-Re(Arg11)CCMSH was labeled with 203Pb in 0.5 M NH4OAc buffer at pH 5.4. The internalization and efflux of 203Pb-DOTA-Re(Arg11)CCMSH were determined in B16/F1 melanoma cells. The pharmacokinetics of 203Pb-DOTA-Re(Arg11)CCMSH were examined in B16/F1 melanoma-bearing C57 mice. A micro-SPECT/CT imaging study was performed with 203Pb-DOTA-Re(Arg11)CCMSH in a B16/F1 melanoma-bearing C57 mouse at 2 h post-injection. Results: 203Pb-DOTA-Re(Arg11)CCMSH was easily prepared in NH4OAc buffer and completely separated from the excess non-radiolabeled peptide by RP-HPLC. 203Pb-DOTA-Re(Arg11)CCMSH displayed fast internalization and extended retention in B16/F1 cells. Approximately 73% of 203Pb-DOTA-Re(Arg11)CCMSH activity internalized after a 20-min incubation at 25C. After incubating the cells in culture media for 20 min, 78% of internalized activity remained in the cells. 203Pb-DOTA-Re(Arg11)CCMSH exhibited similar biodistribution pattern with 212Pb-DOTA-Re(Arg11)CCMSH in B16/F1 melanoma-bearing mice. 203Pb-DOTA-Re(Arg11)CCMSH exhibited the peak tumor uptake of 12.00 +/- 3.20 %ID/g at 1 h post-injection. The tumor uptake gradually decreased to 3.43 +/- 1.12 %ID/g at 48 h post-injection. 203Pb-DOTA-Re(Arg11)CCMSH exhibited the peak tumor to kidney

  16. Melanoma-targeting properties of (99m)technetium-labeled cyclic alpha-melanocyte-stimulating hormone peptide analogues.

    PubMed

    Chen, J; Cheng, Z; Hoffman, T J; Jurisson, S S; Quinn, T P

    2000-10-15

    Preliminary reports have demonstrated that (99m)technetium (Tc)-labeled cyclic [Cys(3,4,10), D-Phe7]alpha-MSH(3-13) (CCMSH) exhibits high tumor uptake and retention values in a murine melanoma mouse model. In this report, the tumor targeting mechanism of 99mTc-CCMSH was studied and compared with four other radiolabeled alpha-melanocyte stimulating hormone (alpha-MSH) peptide analogues: 125I-(Tyr2)-[Nle4, D-Phe7]alpha-MSH [125I-(Tyr2)-NDP]; 99mTc-CGCG-NDP; 99mTc-Gly11-CCMSH; and 99mTc-Nle11-CCMSH. In vitro receptor binding, internalization, and cellular retention of radiolabeled alpha-MSH analogues in B16/F1 murine cell line demonstrated that >70% of the receptor-bound radiolabeled analogues were internalized together with the receptor. Ninety % of the internalized 125I-(Tyr2)-NDP, whereas only 36% of internalized 99mTc-CCMSH, was released from the cells into the medium during a 4-h incubation at 37 degrees C. Two mouse models, C57 mice and severe combined immunodeficient (Scid) mice, inoculated s.c. with B16/F1 murine and TXM-13 human melanoma cells were used for the in vivo studies. Tumor uptake values of 11.32 and 2.39 [% injected dose (ID)/g] for 99mTc-CCMSH at 4 h after injection, resulted in an uptake ratio of tumor:blood of 39.0 and 11.5 in murine melanoma-C57 and human melanoma-Scid mouse models, respectively. Two strategies for decreasing the nonspecific kidney uptake of 99mTc-CCMSH, substitution of Lys11 in CCMSH with Gly11 or Nle11, and lysine coinjection, were evaluated. The biodistribution data for the modified peptides showed that Lys11 replacement dramatically decreased the kidney uptake, whereas the tumor uptakes of 99mTc-Nle11- and 99mTc-Gly11-CCMSH were significantly lower than that of 99mTc-CCMSH. Lysine coinjection significantly decreased the kidney uptake (e.g., from 14.6% ID/g to 4.5% ID/g at 4 h after injection in murine melanoma-C57 mice) without significantly changing the value of tumor uptake of 99mTc-CCMSH. In conclusion, the compact

  17. Interaction of alpha-melanocyte-stimulating hormone, melatonin, cyclic AMP and cyclic GMP in the control of melanogenesis in hair follicle melanocytes in vitro.

    PubMed

    Weatherhead, B; Logan, A

    1981-07-01

    In short-term (48 h) cultures of hair follicles alpha-melanocyte-stimulating hormone (alpha-MSH) and cyclic AMP stimulated melanogenesis through an increase in tyrosinase activity. In contrast cyclic GMP mimicked the effects of melatonin by inhibiting melanin production without causing a concomitant decrease in tyrosinase activity. Both cyclic GMP and melatonin blocked the stimulatory effects of cyclic AMP and alpha-MSH on melanin production but they left the increased levels of tyrosinase activity unaffected. Phosphodiesterase inhibitors (3-isobutyl-1--methylxanthine and papaverine) simultaneously stimulated tyrosinase activity and inhibited melanin production, presumably by allowing endogenous cyclic AMP and cyclic GMP to accumulate intracellularly. It is suggested that whereas MSH stimulates melanogenesis through a cyclic AMP-dependent mechanism there must also be an inhibitory cyclic GMP-dependent mechanism, perhaps activated by melatonin, which operates at some post-tyrosinase step in the melanin biosynthetic pathway. PMID:6267154

  18. Ultraviolet B, melanin and mitochondrial DNA: Photo-damage in human epidermal keratinocytes and melanocytes modulated by alpha-melanocyte-stimulating hormone

    PubMed Central

    Böhm, Markus; Hill, Helene Z.

    2016-01-01

    Alpha-melanocyte-stimulating hormone (alpha-MSH) increases melanogenesis and protects from UV-induced DNA damage. However, its effect on mitochondrial DNA (mtDNA) damage is unknown. We have addressed this issue in a pilot study using human epidermal keratinocytes and melanocytes incubated with alpha-MSH and irradiated with UVB. Real-time touchdown PCR was used to quantify total and deleted mtDNA. The deletion detected encompassed the common deletion but was more sensitive to detection. There were 4.4 times more mtDNA copies in keratinocytes than in melanocytes. Irradiation alone did not affect copy numbers. Alpha-MSH slightly increased copy numbers in both cell types in the absence of UVB and caused a similar small decrease in copy number with dose in both cell types. Deleted copies were nearly twice as frequent in keratinocytes as in melanocytes. Alpha-MSH reduced the frequency of deleted copies by half in keratinocytes but not in melanocytes. UVB dose dependently led to an increase in the deleted copy number in alpha-MSH-treated melanocytes. UVB irradiation had little effect on deleted copy number in alpha-MSH-treated keratinocytes. In summary, alpha-MSH enhances mtDNA damage in melanocytes presumably by increased melanogenesis, while α-MSH is protective in keratinocytes, the more so in the absence of irradiation. PMID:27303631

  19. Ultraviolet B, melanin and mitochondrial DNA: Photo-damage in human epidermal keratinocytes and melanocytes modulated by alpha-melanocyte-stimulating hormone.

    PubMed

    Böhm, Markus; Hill, Helene Z

    2016-01-01

    Alpha-melanocyte-stimulating hormone (alpha-MSH) increases melanogenesis and protects from UV-induced DNA damage. However, its effect on mitochondrial DNA (mtDNA) damage is unknown. We have addressed this issue in a pilot study using human epidermal keratinocytes and melanocytes incubated with alpha-MSH and irradiated with UVB. Real-time touchdown PCR was used to quantify total and deleted mtDNA. The deletion detected encompassed the common deletion but was more sensitive to detection. There were 4.4 times more mtDNA copies in keratinocytes than in melanocytes. Irradiation alone did not affect copy numbers. Alpha-MSH slightly increased copy numbers in both cell types in the absence of UVB and caused a similar small decrease in copy number with dose in both cell types. Deleted copies were nearly twice as frequent in keratinocytes as in melanocytes. Alpha-MSH reduced the frequency of deleted copies by half in keratinocytes but not in melanocytes. UVB dose dependently led to an increase in the deleted copy number in alpha-MSH-treated melanocytes. UVB irradiation had little effect on deleted copy number in alpha-MSH-treated keratinocytes. In summary, alpha-MSH enhances mtDNA damage in melanocytes presumably by increased melanogenesis, while α-MSH is protective in keratinocytes, the more so in the absence of irradiation.

  20. De novo design, synthesis, and pharmacology of alpha-melanocyte stimulating hormone analogues derived from somatostatin by a hybrid approach.

    PubMed

    Han, Guoxia; Haskell-Luevano, Carrie; Kendall, Laura; Bonner, Gregg; Hadley, Mac E; Cone, Roger D; Hruby, Victor J

    2004-03-11

    A number of alpha-melanotropin (alpha-MSH) analogues have been designed de novo, synthesized, and bioassayed at different melanocortin receptors from frog skin (fMC1R) and mouse/rat (mMC1R, rMC3R, mMC4R, and mMC5R). These ligands were designed from somatostatin by a hybrid approach, which utilizes a modified cyclic structure (H-d-Phe-c[Cys---Cys]-Thr-NH(2)) related to somatostatin analogues (e.g. sandostatin) acting at somatostatin receptors, CTAP which binds specifically to micro opioid receptors, and the core pharmacophore of alpha-MSH (His-Phe-Arg-Trp). Ligands designed were H-d-Phe-c[XXX-YYY-ZZZ-Arg-Trp-AAA]-Thr-NH(2) [XXX and AAA = Cys, d-Cys, Hcy, Pen, d-Pen; YYY = His, His(1'-Me), His(3'-Me); ZZZ = Phe and side chain halogen substituted Phe, d-Phe, d-Nal(1'), and d-Nal(2')]. The compounds showed a wide range of bioactivities at the frog skin MC1R; e.g. H-d-Phe-c[Hcy-His-d-Phe-Arg-Trp-Cys]-Thr-NH(2) (6, EC(50) = 0.30 nM) and H-d-Phe-c[Cys-His-d-Phe-Arg-Trp-d-Cys]-Thr-NH(2) (8, EC(50) = 0.10 nM). In addition, when a lactam bridge was used as in H-d-Phe-c[Asp-His-d-Phe-Arg-Trp-Lys]-Thr-NH(2) (7, EC(50) = 0.10 nM), the analogue obtained is as potent as alpha-MSH in the frog skin MC1R assay. Interestingly, switching the bridge of 6 to give H-d-Phe-c[Cys-His-d-Phe-Arg-Trp-Hcy]-Thr-NH(2) (5, EC(50) = 1000 nM) led to a 3000-fold decrease in agonist activity. An increase in steric size in the side chain of d-Phe(7) reduced the bioactivity significantly. For example, H-d-Phe-c[Cys-His-d-Nal(1')-Arg-Trp-d-Cys]-Thr-NH(2) (24) is 2000-fold less active than 9. On the other hand, H-d-Phe-c[Cys-His-d-Phe(p-I)-Arg-Trp-d-Cys]-Thr-NH(2) (23) lost all agonist activity and became a weak antagonist (IC(50) = 1 x 10(-5) M). Furthermore, the modified CTAP analogues with a d-Trp at position 7 all showed weak antagonist activities (EC(50) = 10(-6) to 10(-7) M). Compounds bioassayed at mouse/rat MCRs displayed intriguing results. Most of them are potent at all four receptors tested (m

  1. Nitric oxide enhances the sensitivity of alpaca melanocytes to respond to {alpha}-melanocyte-stimulating hormone by up-regulating melanocortin-1 receptor

    SciTech Connect

    Dong, Yanjun; Cao, Jing; Wang, Haidong; Zhang, Jie; Zhu, Zhiwei; Bai, Rui; Hao, HuanQing; He, Xiaoyan; Fan, Ruiwen; Dong, Changsheng

    2010-06-11

    Nitric oxide (NO) and {alpha}-melanocyte-stimulating hormone ({alpha}-MSH) have been correlated with the synthesis of melanin. The NO-dependent signaling of cellular response to activate the hypothalamopituitary proopiomelanocortin system, thereby enhances the hypophysial secretion of {alpha}-MSH to stimulate {alpha}-MSH-receptor responsive cells. In this study we investigated whether an NO-induced pathway can enhance the ability of the melanocyte to respond to {alpha}-MSH on melanogenesis in alpaca skin melanocytes in vitro. It is important for us to know how to enhance the coat color of alpaca. We set up three groups for experiments using the third passage number of alpaca melanocytes: the control cultures were allowed a total of 5 days growth; the UV group cultures like the control group but the melanocytes were then irradiated everyday (once) with 312 mJ/cm{sup 2} of UVB; the UV + L-NAME group is the same as group UV but has the addition of 300 {mu}M L-NAME (every 6 h). To determine the inhibited effect of NO produce, NO produces were measured. To determine the effect of the NO to the key protein and gene of {alpha}-MSH pathway on melanogenesis, the key gene and protein of the {alpha}-MSH pathway were measured by quantitative real-time PCR and Western immunoblotting. The results provide exciting new evidence that NO can enhance {alpha}-MSH pathway in alpaca skin melanocytes by elevated MC1R. And we suggest that the NO pathway may more rapidly cause the synthesis of melanin in alpaca skin under UV, which at that time elevates the expression of MC1R and stimulates the keratinocytes to secrete {alpha}-MSH to enhance the {alpha}-MSH pathway on melanogenesis. This process will be of considerable interest in future studies.

  2. Nitric oxide enhances the sensitivity of alpaca melanocytes to respond to alpha-melanocyte-stimulating hormone by up-regulating melanocortin-1 receptor.

    PubMed

    Dong, Yanjun; Cao, Jing; Wang, Haidong; Zhang, Jie; Zhu, Zhiwei; Bai, Rui; Hao, HuanQing; He, Xiaoyan; Fan, Ruiwen; Dong, Changsheng

    2010-06-11

    Nitric oxide (NO) and alpha-melanocyte-stimulating hormone (alpha-MSH) have been correlated with the synthesis of melanin. The NO-dependent signaling of cellular response to activate the hypothalamopituitary proopiomelanocortin system, thereby enhances the hypophysial secretion of alpha-MSH to stimulate alpha-MSH-receptor responsive cells. In this study we investigated whether an NO-induced pathway can enhance the ability of the melanocyte to respond to alpha-MSH on melanogenesis in alpaca skin melanocytes in vitro. It is important for us to know how to enhance the coat color of alpaca. We set up three groups for experiments using the third passage number of alpaca melanocytes: the control cultures were allowed a total of 5 days growth; the UV group cultures like the control group but the melanocytes were then irradiated everyday (once) with 312 mJ/cm(2) of UVB; the UV+L-NAME group is the same as group UV but has the addition of 300 microM L-NAME (every 6h). To determine the inhibited effect of NO produce, NO produces were measured. To determine the effect of the NO to the key protein and gene of alpha-MSH pathway on melanogenesis, the key gene and protein of the alpha-MSH pathway were measured by quantitative real-time PCR and Western immunoblotting. The results provide exciting new evidence that NO can enhance alpha-MSH pathway in alpaca skin melanocytes by elevated MC1R. And we suggest that the NO pathway may more rapidly cause the synthesis of melanin in alpaca skin under UV, which at that time elevates the expression of MC1R and stimulates the keratinocytes to secrete alpha-MSH to enhance the alpha-MSH pathway on melanogenesis. This process will be of considerable interest in future studies.

  3. Mapping of the gene encoding the melanocortin-1 ([alpha]-melanocyte stimulating hormone) receptor (MC1R) to human chromosome 16q24. 3 by fluorescence in situ hybridization

    SciTech Connect

    Gantz, I.; Yamada, Tadataka; Tashiro, Takao; Konda, Yoshitaka; Shimoto, Yoshimasa; Miwa, Hiroto; Trent, J.M. )

    1994-01-15

    [alpha]-Melanocyte stimulating hormone ([alpha]-MSH), a hormone originally named for its ability to regulate pigmentation of melanocytes, is a 13-amino-acid post-translational product of the pro-opiomelanocortin (POMC) gene. [alpha]-MSH and the other products of POMC processing, which share the core heptapeptide amino acid sequence Met-Glu (Gly)-His-Phe-Arg-Trp-Gly (Asp), the adrenocorticotropic hormone (ACTH), [beta]-MSH, and [gamma]-MSH, are collectively referred to as melanocortins. While best known for their effects on the melanocyte (pigmentation) and adrenal cortical cells (steroidogenesis), melanocortins have been postulated to function in diverse activities, including enhancement of learning and memory, control of the cardiovascular system, analgesia, thermoregulation, immunomodulation, parturition, and neurotrophism. To identify the chromosomal band encoding the human melanocortin-1 receptor gene, 1 [mu]g of an EMBL clone coding region of the human MC1R and approximately 15 kb of surrounding DNA was labeled with biotin and hybridized to human metaphase chromosomes as previously described. The results indicate that the human MC1R gene is localized to 16q24.3. 15 refs., 1 fig.

  4. β-Lactam Antibiotics Renaissance

    PubMed Central

    Qin, Wenling; Panunzio, Mauro; Biondi, Stefano

    2014-01-01

    Since the 1940s β-lactam antibiotics have been used to treat bacterial infections. However, emergence and dissemination of β-lactam resistance has reached the point where many marketed β-lactams no longer are clinically effective. The increasing prevalence of multidrug-resistant bacteria and the progressive withdrawal of pharmaceutical companies from antibiotic research have evoked a strong reaction from health authorities, who have implemented initiatives to encourage the discovery of new antibacterials. Despite this gloomy scenario, several novel β-lactam antibiotics and β-lactamase inhibitors have recently progressed into clinical trials, and many more such compounds are being investigated. Here we seek to provide highlights of recent developments relating to the discovery of novel β-lactam antibiotics and β-lactamase inhibitors. PMID:27025744

  5. Overcoming Resistance to β-Lactam Antibiotics

    PubMed Central

    Worthington, Roberta J.; Melander, Christian

    2013-01-01

    β-Lactam antibiotics are one of the most important antibiotic classes but are plagued by problems of resistance and the development of new β-lactam antibiotics through side chain modification of existing β-lactam classes is not keeping pace with resistance development. In this perspective we summarize small molecule strategies to overcome resistance to β-lactam antibiotics. These approaches include the development of β-lactamase inhibitors and compounds that interfere with the ability of the bacteria to sense an antibiotic threat and activate their resistance mechanisms. PMID:23530949

  6. Process for the production of lactams

    SciTech Connect

    Matson, M.S.

    1989-01-24

    A process is described for producing a lactam comprising contacting a feedstock selected from the group consisting of acid anhydrides, dicarboxylic acids, lactones, and imides, the feedstock having 4 to 6 carbon atoms in the backbone with a catalyst system comprising palladium and at least one second metal selected from the group consisting of ruthenium, rhodium and rhenium under conditions to convert at least a portion of the feedstock to the lactam.

  7. Resistance to beta-lactams--the permutations.

    PubMed

    Amyes, S G B

    2003-12-01

    The beta-lactam family of antimicrobials, in particular penicillins and cephalosporins, is the mainstay of treatment for community-acquired infections. However, the emergence of resistant isolates to these agents has raised concerns regarding the continued efficacy of existing therapies. Resistance to beta-lactams is most commonly expressed by the microbial production of beta-lactamases that hydrolyze the beta-lactam ring. Three further resistance mechanisms include conformational changes in penicillin-binding proteins (PBPs); permeability changes in the outer membrane; and active efflux of the antimicrobial. In addition to the pre-requisite efficacy and tolerability profiles, new beta-lactams should address these four resistance mechanisms. Overcoming resistance may be a serendipitous event or arrived at by design. A unique synthetic beta-lactam class, which demonstrates promise in terms of its activity against the range of bacteria responsible for community-acquired infections and its inherent stability to hydrolysis by beta-lactamases, is the penems. This discrete class of hybrid molecules combines properties from the penicillin (penam) and cephalosporin (cephem) beta-lactam classes. Faropenem is an example of a penem with a broad spectrum of activity designed to address these resistance issues. PMID:14998075

  8. Lactam inhibiting Streptococcus mutans growth on titanium.

    PubMed

    Xavier, J G; Geremias, T C; Montero, J F D; Vahey, B R; Benfatti, C A M; Souza, J C M; Magini, R S; Pimenta, A L

    2016-11-01

    The aim of this work was to analyze the activity of novel synthetic lactams on preventing biofilm formation on titanium surfaces. Titanium (Ti6Al4V) samples were exposed to Streptococcus mutans cultures in the presence or absence of a synthetic lactam. After 48h incubation, planktonic growth was determined by spectrophotometry. Biofilm was evaluated by crystal violet staining and colony forming units (CFU·ml(-)(1)), followed by scanning electron microscopy (SEM). Results showed that the average of adhered viable cells was approximately 1.5×10(2)CFU/ml in the presence of lactam and 4×10(2)CFU/ml in its absence. This novel compound was considerable active in reducing biofilm formation over titanium surfaces, indicating its potential for the development of antimicrobial drugs targeting the inhibition of the initial stages of bacterial biofilms on dental implants abutments. PMID:27524086

  9. β-lactam-associated eosinophilic colitis.

    PubMed

    Mogilevski, Tamara; Nickless, David; Hume, Sam

    2015-01-01

    A 42-year-old man with a history of childhood asthma presented with a 2-week history of watery diarrhoea and marked peripheral eosinophilia in the setting of recent use of cephalexin. His colonoscopy revealed patchy colitis. Biopsies were consistent with eosinophilic colitis. Two months later he received a course of amoxicillin resulting in recurrence of peripheral eosinophilia. Given the time-frame of β-lactam administration to symptom onset and elimination of all other precipitating causes, he was diagnosed with β-lactam-associated eosinophilic colitis. The patient's symptoms resolved and peripheral eosinophil count decreased with no specific treatment. Eosinophilic colitis is a rare heterogeneous condition, the pathogenesis of which is likely to be an interplay between environmental and genetic factors. It can be secondary to a helminthic infection or a drug reaction and has been associated with ulcerative colitis. If secondary causes of eosinophilic colitis have been excluded, the mainstay of treatment is with corticosteroids. PMID:26106168

  10. Sensor histidine kinase is a β-lactam receptor and induces resistance to β-lactam antibiotics.

    PubMed

    Li, Lu; Wang, Qiyao; Zhang, Hui; Yang, Minjun; Khan, Mazhar I; Zhou, Xiaohui

    2016-02-01

    β-Lactams disrupt bacterial cell wall synthesis, and these agents are the most widely used antibiotics. One of the principle mechanisms by which bacteria resist the action of β-lactams is by producing β-lactamases, enzymes that degrade β-lactams. In Gram-negative bacteria, production of β-lactamases is often induced in response to the antibiotic-associated damage to the cell wall. Here, we have identified a previously unidentified mechanism that governs β-lactamase production. In the Gram-negative enteric pathogen Vibrio parahaemolyticus, we found a histidine kinase/response regulator pair (VbrK/VbrR) that controls expression of a β-lactamase. Mutants lacking either VbrK or VbrR do not produce the β-lactamase and are no longer resistant to β-lactam antibiotics. Notably, VbrK autophosphorylation is activated by β-lactam antibiotics, but not by other lactams. However, single amino acid substitutions in the putative periplasmic binding pocket of VbrK leads its phosphorylation in response to both β-lactam and other lactams, suggesting that this kinase is a β-lactam receptor that can directly detect β-lactam antibiotics instead of detecting the damage to cell wall resulting from β-lactams. In strong support of this idea, we found that purified periplasmic sensor domain of VbrK binds penicillin, and that such binding is critical for VbrK autophosphorylation and β-lactamase production. Direct recognition of β-lactam antibiotics by a histidine kinase receptor may represent an evolutionarily favorable mechanism to defend against β-lactam antibiotics.

  11. Biobased lactams as novel arthropod repellents.

    PubMed

    Chauhan, Kamlesh R; Khanna, Hemant; Bathini, Nagendra Babu; Le, Thanh C; Grieco, John

    2014-12-01

    Enanatiomerically pure 4aS,7S,7aR and 4aS,7S,7aS-nepetalactams and their analogs have been prepared in just two steps from 4aS,7S,7aR and 4aS,7S,7aS-nepetalactones, major components of catnip oil. Lactams or cyclic amides from iridoid monoterpenes are generated and being evaluated as a new class of compounds as arthropod deterrents against disease vectors. PMID:25632454

  12. Ureidopenicillins and beta-lactam/beta-lactamase inhibitor combinations.

    PubMed

    Bush, L M; Johnson, C C

    2000-06-01

    Although research and development of new penicillins have declined, penicillins continue to be essential antibiotics for the treatment and prophylaxis of infectious diseases. The most recent additions are the ureidopenicillins and beta-lactam/beta-lactamase inhibitor combinations. This article reviews the spectrum of activity, toxicity, pharmacokinetics, and clinical uses of the ureidopenicillins, and the beta-lactam/beta-lactamase inhibitor combination agents.

  13. The antioxidant effect of derivatives pyroglutamic lactam

    SciTech Connect

    Rohadi, Atisya; Lazim, Azwani Mat; Hasbullah, Siti Aishah

    2013-11-27

    Diphenylpicrylhydrazyl (DPPH) is widely used for quickly accessing the ability of polyphenols to transfer labile H atoms to radicals. The antioxidant activity of all the synthesized compounds was screened by DPPH method. Compound (4) showed 54% antioxidant potential while all other compounds were found to have moderate to have moderate to mild antioxidant activity ranging from 47–52%. Pyroglutamic lactams have been synthesized stereoselectively in racemic form from levulinic acid as bifunctional adduct using convertible isocyanide in one-pot Ugi 4-center-3-component condensation reaction (U-4C-3CR). The product formed provides biologically interesting products in excellent yields in a short reaction time. The structures of the synthesized compounds were elucidated using spectroscopic data and elemental analysis.

  14. The antioxidant effect of derivatives pyroglutamic lactam

    NASA Astrophysics Data System (ADS)

    Rohadi, Atisya; Lazim, Azwani Mat; Hasbullah, Siti Aishah

    2013-11-01

    Diphenylpicrylhydrazyl (DPPH) is widely used for quickly accessing the ability of polyphenols to transfer labile H atoms to radicals. The antioxidant activity of all the synthesized compounds was screened by DPPH method. Compound (4) showed 54% antioxidant potential while all other compounds were found to have moderate to have moderate to mild antioxidant activity ranging from 47-52%. Pyroglutamic lactams have been synthesized stereoselectively in racemic form from levulinic acid as bifunctional adduct using convertible isocyanide in one-pot Ugi 4-center-3-component condensation reaction (U-4C-3CR). The product formed provides biologically interesting products in excellent yields in a short reaction time. The structures of the synthesized compounds were elucidated using spectroscopic data and elemental analysis.

  15. Endopeptidase-Mediated Beta Lactam Tolerance

    PubMed Central

    Dörr, Tobias; Davis, Brigid M.; Waldor, Matthew K.

    2015-01-01

    In many bacteria, inhibition of cell wall synthesis leads to cell death and lysis. The pathways and enzymes that mediate cell lysis after exposure to cell wall-acting antibiotics (e.g. beta lactams) are incompletely understood, but the activities of enzymes that degrade the cell wall (‘autolysins’) are thought to be critical. Here, we report that Vibrio cholerae, the cholera pathogen, is tolerant to antibiotics targeting cell wall synthesis. In response to a wide variety of cell wall- acting antibiotics, this pathogen loses its rod shape, indicative of cell wall degradation, and becomes spherical. Genetic analyses revealed that paradoxically, V. cholerae survival via sphere formation required the activity of D,D endopeptidases, enzymes that cleave the cell wall. Other autolysins proved dispensable for this process. Our findings suggest the enzymes that mediate cell wall degradation are critical for determining bacterial cell fate - sphere formation vs. lysis – after treatment with antibiotics that target cell wall synthesis. PMID:25884840

  16. The road to avibactam: the first clinically useful non-β-lactam working somewhat like a β-lactam.

    PubMed

    Wang, David Yuxin; Abboud, Martine I; Markoulides, Marios S; Brem, Jürgen; Schofield, Christopher J

    2016-06-01

    Avibactam, which is the first non-β-lactam β-lactamase inhibitor to be introduced for clinical use, is a broad-spectrum serine β-lactamase inhibitor with activity against class A, class C, and, some, class D β-lactamases. We provide an overview of efforts, which extend to the period soon after the discovery of the penicillins, to develop clinically useful non-β-lactam compounds as antibacterials, and, subsequently, penicillin-binding protein and β-lactamase inhibitors. Like the β-lactam inhibitors, avibactam works via a mechanism involving covalent modification of a catalytically important nucleophilic serine residue. However, unlike the β-lactam inhibitors, avibactam reacts reversibly with its β-lactamase targets. We discuss chemical factors that may account for the apparently special nature of β-lactams and related compounds as antibacterials and β-lactamase inhibitors, including with respect to resistance. Avenues for future research including non-β-lactam antibacterials acting similarly to β-lactams are discussed.

  17. The road to avibactam: the first clinically useful non-β-lactam working somewhat like a β-lactam.

    PubMed

    Wang, David Yuxin; Abboud, Martine I; Markoulides, Marios S; Brem, Jürgen; Schofield, Christopher J

    2016-06-01

    Avibactam, which is the first non-β-lactam β-lactamase inhibitor to be introduced for clinical use, is a broad-spectrum serine β-lactamase inhibitor with activity against class A, class C, and, some, class D β-lactamases. We provide an overview of efforts, which extend to the period soon after the discovery of the penicillins, to develop clinically useful non-β-lactam compounds as antibacterials, and, subsequently, penicillin-binding protein and β-lactamase inhibitors. Like the β-lactam inhibitors, avibactam works via a mechanism involving covalent modification of a catalytically important nucleophilic serine residue. However, unlike the β-lactam inhibitors, avibactam reacts reversibly with its β-lactamase targets. We discuss chemical factors that may account for the apparently special nature of β-lactams and related compounds as antibacterials and β-lactamase inhibitors, including with respect to resistance. Avenues for future research including non-β-lactam antibacterials acting similarly to β-lactams are discussed. PMID:27327972

  18. Synthesis of quaternary α-perfluoroalkyl lactams via electrophilic perfluoroalkylation.

    PubMed

    Katayev, D; Václavík, J; Brüning, F; Commare, B; Togni, A

    2016-03-14

    Efficient protocols enabling the rapid installation of trifluoromethyl, as well as further functionalized fluoroalkyl groups by an electrophilic perfluoroalkylation of lactam-derived ketene silyl amides (KSAs) using hypervalent iodine reagents 1 and 2 have been developed. PMID:26893124

  19. Recent Approaches Toward Solid Phase Synthesis of β-Lactams

    NASA Astrophysics Data System (ADS)

    Mandal, Bablee; Ghosh, Pranab; Basu, Basudeb

    Since the discovery of penicillin in 1929, β-lactam antibiotics have been recognized as potentially chemotherapeutic drugs of incomparable effectiveness, conjugating a broad spectrum of activity with very low toxicity. The primary motif azetidin-2-one ring (β-lactam) has been considered as specific pharmacophores and scaffolds. With the advent of combinatorial chemistry and automated parallel synthesis coupled with ample interests from the pharmaceutical industries, recent trends have been driven mostly by adopting solid phase techniques and polymer-supported synthesis of β-lactams. The present survey will present an overview of the developments on the polymer-supported and solid phase techniques for the preparation of β-lactam ring or β-lactam containing antibiotics published over the last decade. Both unsubstituted and substitutions with different functional groups at various positions of β-lactams have been synthesized using solid phase technology. However, Wang resin and application of Staudinger [2+2] cycloaddition reaction have remained hitherto the major choice. It may be expected that other solid phase approaches involving different resins would be developed in the coming years.

  20. Penicillin and beta-lactam allergy: epidemiology and diagnosis.

    PubMed

    Macy, Eric

    2014-11-01

    Penicillin is the most common beta-lactam antibiotic allergy and the most common drug class allergy, reported in about 8% of individuals using health care in the USA. Only about 1% of individuals using health care in the USA have a cephalosporin allergy noted in their medical record, and other specific non-penicillin, non-cephalosporin beta-lactam allergies are even rarer. Most reported penicillin allergy is not associated with clinically significant IgE-mediated reactions after penicillin rechallenge. Un-verified penicillin allergy is a significant and growing public health problem. Clinically significant IgE-mediated penicillin allergy can be safely confirmed or refuted using skin testing with penicilloyl-poly-lysine and native penicillin G and, if skin test is negative, an oral amoxicillin challenge. Acute tolerance of an oral therapeutic dose of a penicillin class antibiotic is the current gold standard test for a lack of clinically significant IgE-mediated penicillin allergy. Cephalosporins and other non-penicillin beta-lactams are widely, safely, and appropriately used in individuals, even with confirmed penicillin allergy. There is little, if any, clinically significant immunologic cross-reactivity between penicillins and other beta-lactams. Routine cephalosporin skin testing should be restricted to research settings. It is rarely needed clinically to safely manage patients and has unclear predictive value at this time. The use of alternative cephalosporins, with different side chains, is acceptable in the setting of a specific cephalosporin allergy. Carbapenems and monobactams are also safely used in individuals with confirmed penicillin allergy. A certain predictable, but low, rate of adverse reactions will occur with all beta-lactam antibiotic use both pre- and post-beta-lactam allergy evaluations.

  1. Management of allergy to penicillins and other beta-lactams.

    PubMed

    Mirakian, R; Leech, S C; Krishna, M T; Richter, A G; Huber, P A J; Farooque, S; Khan, N; Pirmohamed, M; Clark, A T; Nasser, S M

    2015-02-01

    The Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI) and an expert panel have prepared this guidance for the management of immediate and non-immediate allergic reactions to penicillins and other beta-lactams. The guideline is intended for UK specialists in both adult and paediatric allergy and for other clinicians practising allergy in secondary and tertiary care. The recommendations are evidence based, but where evidence is lacking, the panel reached consensus. During the development of the guideline, all BSACI members were consulted using a Web-based process and all comments carefully considered. Included in the guideline are epidemiology of allergic reactions to beta-lactams, molecular structure, formulations available in the UK and a description of known beta-lactam antigenic determinants. Sections on the value and limitations of clinical history, skin testing and laboratory investigations for both penicillins and cephalosporins are included. Cross-reactivity between penicillins and cephalosporins is discussed in detail. Recommendations on oral provocation and desensitization procedures have been made. Guidance for beta-lactam allergy in children is given in a separate section. An algorithm to help the clinician in the diagnosis of patients with a history of penicillin allergy has also been included.

  2. Spirocyclic β-Lactams: Synthesis and Biological Evaluation of Novel Heterocycles

    NASA Astrophysics Data System (ADS)

    Bari, Shamsher S.; Bhalla, Aman

    β-Lactam rings containing compounds are a group of antibiotics of unparalleled importance in chemotherapy. Considerable effort has been reported in the development of novel, more effective β-lactam compounds as well as their biological evaluation. This article reviews the progress made in the stereoselective synthesis of spiro-β-lactams, a unique class of heterocycles, their biological evaluation, and their applications in various related fields. The introductory paragraph highlights the significance of the β-lactam chemistry and is followed by an overview of monocyclic-, bicyclic-, and tricyclic-β-lactams. The other sections of the article deal with the stereoselective synthesis and biological evaluation of spiro-β-lactams, including their use as synthetic intermediates for β-turn mimics and β-turn nucleators. The potential of spiro-β-lactams as cholesterol absorption inhibitors, β-lactamase inhibitors, and antiviral, antibacterial, and antimicrobial agents has also been described.

  3. Modulating the hydration behaviour of calcium chloride by lactam complexation.

    PubMed

    Perrin, Andrea; Musa, Osama M; Steed, Jonathan W

    2016-07-26

    Complexation of calcium chloride with bis(lactam) ligand L1 allows the formation of both an unstable anhydrous complex, an aqua complex {[Ca2(μ-L1)2(H2O)9]Cl4]}n (1) and a related hydrate incorporating additional lattice water of crystallization {[Ca(μ-L1)(H2O)5]Cl2·H2O}n (2). Related mono(lactam) L2 does not form aqua complexes but the anhydrous complex {[CaCl2(μ-L2)2]}n (3), is highly deliquescent. An unusual ethanol solvate is also reported {[CaCl2(L2)(EtOH)]}n (4).

  4. Modulating the hydration behaviour of calcium chloride by lactam complexation.

    PubMed

    Perrin, Andrea; Musa, Osama M; Steed, Jonathan W

    2016-07-26

    Complexation of calcium chloride with bis(lactam) ligand L1 allows the formation of both an unstable anhydrous complex, an aqua complex {[Ca2(μ-L1)2(H2O)9]Cl4]}n (1) and a related hydrate incorporating additional lattice water of crystallization {[Ca(μ-L1)(H2O)5]Cl2·H2O}n (2). Related mono(lactam) L2 does not form aqua complexes but the anhydrous complex {[CaCl2(μ-L2)2]}n (3), is highly deliquescent. An unusual ethanol solvate is also reported {[CaCl2(L2)(EtOH)]}n (4). PMID:27411017

  5. [ACUTE RESPIRATORY DISTRESS SYNDROME CAUSED BY BETA-LACTAM ANTIBIOTICS].

    PubMed

    Skipskiy, I M; Efimov, N W; Remizov, A S; Miroshnikov, B I

    2015-01-01

    We report a case of iatrogenic lesion of the lungs in a 61 year old patient following simultaneous subtotal resection and plastic surgery of esophagus using a gastric stem with the removed cardial portion along with subtotal mediastinal lympho-dissection for the treatment of 2 x 1 cm T2NoMO tumour in the middle third of esophagus. The patient's medical history contained no evidence of previous pulmonary pathology, preoperative chest X-ray study revealed neither focal nor infiltrative changes in the lungs. On day 5 after extensive surgical intervention with the use of a parenteral beta-lactam antibiotic, the patient developed acute respiratory distress the symptoms of which increased wavelike during the next 4.5 weeks in association with subfebrility, leukocytosis, enhanced ESR and large shaded areas in the lungs. These conditions were regarded as signs of pneumonia that required the application of increasingly more powerful beta-lactam antibiotics. All these symptoms became less apparent and completely disappeared within 2 days and 1 week respectively after withdrawal of the antibiotics and prescription of parenteral prednisolone. It confirmed the iatrogenic origin of lung lesions regarded by the authors as recurring acute respiratory distress syndrome caused by beta-lactam antibiotics.

  6. A Deep Cavitand Templates Lactam Formation in Water.

    PubMed

    Mosca, Simone; Yu, Yang; Gavette, Jesse V; Zhang, Kang-Da; Rebek, Julius

    2015-11-25

    Cyclization reactions are common processes in organic chemistry and show familiar patterns of reaction rates vs ring size. While the details vary with the nature of bond being made and the number of unsaturated atoms, small rings typically form quickly despite angle strain, medium size rings form very slowly due to internal strains, and large rings form slowly (when they form at all) because fewer and less probable conformations bring the ends of the substrate together. High dilution is commonly used to slow the competing bi- and higher molecular processes. Here we apply cavitands to the formation of medium size lactams from ω-amino acids in aqueous (D2O) solution. The cavitands bind the amino acids in folded conformations that favor cyclization by bringing the ends closer together. Yields of a 12-membered lactam are improved 4.1-fold and 13-membered lactam 2.8-fold by the cavitand template. The results open possibilities for moving organic reactions into water even when the processes involve dehydration. PMID:26540097

  7. β-Lactam Resistance Mechanisms: Gram-Positive Bacteria and Mycobacterium tuberculosis.

    PubMed

    Fisher, Jed F; Mobashery, Shahriar

    2016-01-01

    The value of the β-lactam antibiotics for the control of bacterial infection has eroded with time. Three Gram-positive human pathogens that were once routinely susceptible to β-lactam chemotherapy-Streptococcus pneumoniae, Enterococcus faecium, and Staphylococcus aureus-now are not. Although a fourth bacterium, the acid-fast (but not Gram-positive-staining) Mycobacterium tuberculosis, has intrinsic resistance to earlier β-lactams, the emergence of strains of this bacterium resistant to virtually all other antibiotics has compelled the evaluation of newer β-lactam combinations as possible contributors to the multidrug chemotherapy required to control tubercular infection. The emerging molecular-level understanding of these resistance mechanisms used by these four bacteria provides the conceptual framework for bringing forward new β-lactams, and new β-lactam strategies, for the future control of their infections. PMID:27091943

  8. Secondary metabolism in simulated microgravity: beta-lactam production by Streptomyces clavuligerus

    NASA Technical Reports Server (NTRS)

    Fang, A.; Pierson, D. L.; Mishra, S. K.; Koenig, D. W.; Demain, A. L.

    1997-01-01

    Rotating bioreactors designed at NASA's Johnson Space Center were used to simulate a microgravity environment in which to study secondary metabolism. The system examined was beta-lactam antibiotic production by Streptomyces clavuligerus. Both growth and beta-lactam production occurred in simulated microgravity. Stimulatory effects of phosphate and L-lysine, previously detected in normal gravity, also occurred in simulated microgravity. The degree of beta-lactam antibiotic production was markedly inhibited by simulated microgravity.

  9. Involvement of a 43-kilodalton outer membrane protein in beta-lactam resistance of Shigella dysenteriae.

    PubMed Central

    Kar, A K; Ghosh, A S; Chauhan, K; Ahamed, J; Basu, J; Chakrabarti, P; Kundu, M

    1997-01-01

    A beta-lactam-sensitive strain (C152) of Shigella dysenteriae showed two major outer membrane proteins (OMPs) with M(r)s of 43,000 and 38,000, while the clinical isolate M2 lacked the 43,000-Mr OMP, which acted as a channel for beta-lactam antibiotics. Permeability of beta-lactams across the outer membrane (OM) of M2 was lower than that across the OM of C152. Mutants deficient in the 43-kDa OMP could be selected in vitro from strain C152 in the presence of cefoxitin. All beta-lactam-resistant strains were sensitive to imipenem. PMID:9333070

  10. Molecular Regulation of β-Lactam Biosynthesis in Filamentous Fungi

    PubMed Central

    Brakhage, Axel A.

    1998-01-01

    The most commonly used β-lactam antibiotics for the therapy of infectious diseases are penicillin and cephalosporin. Penicillin is produced as an end product by some fungi, most notably by Aspergillus (Emericella) nidulans and Penicillium chrysogenum. Cephalosporins are synthesized by both bacteria and fungi, e.g., by the fungus Acremonium chrysogenum (Cephalosporium acremonium). The biosynthetic pathways leading to both secondary metabolites start from the same three amino acid precursors and have the first two enzymatic reactions in common. Penicillin biosynthesis is catalyzed by three enzymes encoded by acvA (pcbAB), ipnA (pcbC), and aatA (penDE). The genes are organized into a cluster. In A. chrysogenum, in addition to acvA and ipnA, a second cluster contains the genes encoding enzymes that catalyze the reactions of the later steps of the cephalosporin pathway (cefEF and cefG). Within the last few years, several studies have indicated that the fungal β-lactam biosynthesis genes are controlled by a complex regulatory network, e.g., by the ambient pH, carbon source, and amino acids. A comparison with the regulatory mechanisms (regulatory proteins and DNA elements) involved in the regulation of genes of primary metabolism in lower eukaryotes is thus of great interest. This has already led to the elucidation of new regulatory mechanisms. Furthermore, such investigations have contributed to the elucidation of signals leading to the production of β-lactams and their physiological meaning for the producing fungi, and they can be expected to have a major impact on rational strain improvement programs. The knowledge of biosynthesis genes has already been used to produce new compounds. PMID:9729600

  11. γ-Lactam alkaloids from the flower buds of daylily.

    PubMed

    Matsumoto, Takahiro; Nakamura, Seikou; Nakashima, Souichi; Ohta, Tomoe; Yano, Mamiko; Tsujihata, Junichiro; Tsukioka, Junko; Ogawa, Keiko; Fukaya, Masashi; Yoshikawa, Masayuki; Matsuda, Hisashi

    2016-07-01

    Four new alkaloids, hemerocallisamines IV-VII, were isolated from the methanol extract of flower buds of daylily. The chemical structures of the new compounds were elucidated on the basis of chemical and physicochemical evidence. The absolute stereochemistry of the hemerocallisamines IV-VI was elucidated by the application of the modified Mosher's method, HPLC analysis, and optical rotation. In the present study, the isolated alkaloids significantly inhibited the aggregation of Aβ42 in vitro. This is the first report about bioactive alkaloids with a γ-lactam ring from daylily. In addition, isolated nucleosides showed accelerative effects on neurite outgrowth under the non-fasting condition. PMID:26849229

  12. γ-Lactam alkaloids from the flower buds of daylily.

    PubMed

    Matsumoto, Takahiro; Nakamura, Seikou; Nakashima, Souichi; Ohta, Tomoe; Yano, Mamiko; Tsujihata, Junichiro; Tsukioka, Junko; Ogawa, Keiko; Fukaya, Masashi; Yoshikawa, Masayuki; Matsuda, Hisashi

    2016-07-01

    Four new alkaloids, hemerocallisamines IV-VII, were isolated from the methanol extract of flower buds of daylily. The chemical structures of the new compounds were elucidated on the basis of chemical and physicochemical evidence. The absolute stereochemistry of the hemerocallisamines IV-VI was elucidated by the application of the modified Mosher's method, HPLC analysis, and optical rotation. In the present study, the isolated alkaloids significantly inhibited the aggregation of Aβ42 in vitro. This is the first report about bioactive alkaloids with a γ-lactam ring from daylily. In addition, isolated nucleosides showed accelerative effects on neurite outgrowth under the non-fasting condition.

  13. Discovery of Potent Carbonic Anhydrase and Acetylcholinesterase Inhibitors: 2-Aminoindan β-Lactam Derivatives

    PubMed Central

    Genç, Hayriye; Kalin, Ramazan; Köksal, Zeynep; Sadeghian, Nastaran; Kocyigit, Umit M.; Zengin, Mustafa; Gülçin, İlhami; Özdemir, Hasan

    2016-01-01

    β-Lactams are pharmacologically important compounds because of their various biological uses, including antibiotic and so on. β-Lactams were synthesized from benzylidene-inden derivatives and acetoxyacetyl chloride. The inhibitory effect of these compounds was examined for human carbonic anhydrase I and II (hCA I, and II) and acetylcholinesterase (AChE). The results reveal that β-lactams are inhibitors of hCA I, II and AChE. The Ki values of β-lactams (2a–k) were 0.44–6.29 nM against hCA I, 0.93–8.34 nM against hCA II, and 0.25–1.13 nM against AChE. Our findings indicate that β-lactams (2a–k) inhibit both carbonic anhydrases (CA) isoenzymes and AChE at low nanomolar concentrations. PMID:27775608

  14. β-lactams against emerging 'superbugs': progress and pitfalls.

    PubMed

    Skalweit Helfand, Marion

    2008-07-01

    Bacterial resistance to antimicrobial agents is an ever-growing problem. So-called 'superbugs', such as multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa harboring multiple resistance determinants, including extended-spectrum β-lactamases, carbapenemases, efflux pumps and downregulated outer-membrane proteins or porins, are becoming more prevalent in hospital, intensive and long-term care settings. Enterobacteriaceae are also acquiring a myriad of β-lactamases, such as class A and D carbapenemases, and plasmid-borne class C cephalosporinases. Gram-positive superbugs, including methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-intermediate or heteroglycopeptide-intermediate S. aureus, vancomycin-resistant S. aureus and penicillin-resistant Streptococcus pneumoniae (PRSP), are problematic pathogens, both in the hospital and in the community (e.g., community-acquired MRSA and PRSP). β-lactam antibiotics remain among the most effective and safest anti-infectives in use, although their utility is being severely challenged by these superbugs. This review will discuss aspects of resistance seen in these pathogens and will review some of the newer β-lactam agents, both investigational and in clinical use, that target these superbugs.

  15. Developments in liquid membrane separation of beta-lactam antibiotics.

    PubMed

    Ghosh, A C; Bora, M M; Dutta, N N

    1996-04-01

    This paper presents an overview on the developments in liquid membrane separation and purification of commercially important beta-lactam antibiotics. Reactive extraction via liquid-liquid ion exchange or ion-pair extraction mechanism can be exploited to develop liquid membrane processes for separation and concentration of penicillins and cephalosporins. Because of high selectivity and flux, liquid membrane processes can be adopted for direct extraction of beta-lactams from fermentation broth. Other advantages of liquid membrane technologies are low capital and operating costs, compact unit installation in commercial plants, low material inventory, etc. Both emulsion liquid membrane and supported liquid membrane techniques can be effective under the reactive extraction conditions. However, the stability problems of liquid membrane should be resolved before commercial application can be established. Alternately, reactive extraction in non-dispersive mode with hollow fiber membranes can be an attractive and viable strategy for practical application. Applicability of the liquid membrane processes has been discussed from process engineering and design considerations.

  16. 76 FR 14024 - Draft Guidance for Industry on Non-Penicillin Beta-Lactam Risk Assessment: A CGMP Framework...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-15

    ... HUMAN SERVICES Food and Drug Administration Draft Guidance for Industry on Non-Penicillin Beta-Lactam... guidance for industry entitled ``Non-Penicillin Beta-Lactam Risk Assessment: A CGMP Framework.'' This... non- penicillin beta-lactam antibiotics. The draft guidance is intended to assist manufacturers...

  17. Comparative Treatment Failure Rates of Respiratory Fluoroquinolones or β-Lactam + Macrolide Versus β-Lactam Alone in the Treatment for Community-Acquired Pneumonia in Adult Outpatients

    PubMed Central

    Lee, Meng-Tse Gabriel; Lee, Shih-Hao; Chang, Shy-Shin; Chan, Ya-Lan; Pang, Laura; Hsu, Sue-Ming; Lee, Chien-Chang

    2015-01-01

    Abstract No comparative effectiveness study has been conducted for the following 3 antibiotics: respiratory fluoroquinolone, β-lactam, and β-lactam + advanced macrolide. To gain insights into the real-world clinical effectiveness of these antibiotics for community-acquired pneumonia in adult outpatients, our study investigated the treatment failure rates in 2 million representative participants from the National Health Informatics Project (NHIP) of Taiwan. A new-user cohort design was used to follow NHIP participants from January 2000 until December 2009. Treatment failure was defined by either one of the following events: a second antibiotic prescription, hospitalization due to CAP, an emergency department visit with a diagnosis of CAP, or 30-day nonaccident-related mortality. From 2006 to 2009, we identified 9256 newly diagnosed CAP outpatients, 1602 of whom were prescribed levofloxacin, 2100 were prescribed moxifloxacin, 5049 were prescribed β-lactam alone, and 505 were prescribed advanced macrolide + β-lactam. Compared with the β-lactam-based regimen, the propensity score-matched odds ratio for composite treatment failure was 0.81 (95% CI, 0.67–0.97) for moxifloxacin, 1.10 (95% CI, 0.90–1.35) for levofloxacin, and 0.95 (95% CI, 0.67–1.35) for macrolide +β-lactam. Moxifloxacin was associated with lower treatment failure rates compared with β-lactam alone, or levofloxacin in Taiwanese CAP outpatients. However, due to inherent limitations in our claims database, more randomized controlled trials are required before coming to a conclusion on which antibiotic is more effective for Taiwanese CAP outpatients. More population-based comparative effectiveness studies are also encouraged and should be considered as an integral piece of evidence in local CAP treatment guidelines. PMID:26426664

  18. Carbamoyl Radical-Mediated Synthesis and Semipinacol Rearrangement of β-Lactam Diols

    PubMed Central

    Betou, Marie; Male, Louise; Steed, Jonathan W; Grainger, Richard S

    2014-01-01

    In an approach to the biologically important 6-azabicyclo[3.2.1]octane ring system, the scope of the tandem 4-exo-trig carbamoyl radical cyclization—dithiocarbamate group transfer reaction to ring-fused β-lactams is evaluated. β-Lactams fused to five-, six-, and seven-membered rings are prepared in good to excellent yield, and with moderate to complete control at the newly formed dithiocarbamate stereocentre. No cyclization is observed with an additional methyl substituent on the terminus of the double bond. Elimination of the dithiocarbamate group gives α,β- or β,γ-unsaturated lactams depending on both the methodology employed (base-mediated or thermal) and the nature of the carbocycle fused to the β-lactam. Fused β-lactam diols, obtained from catalytic OsO4-mediated dihydroxylation of α,β-unsaturated β-lactams, undergo semipinacol rearrangement via the corresponding cyclic sulfite or phosphorane to give keto-bridged bicyclic amides by exclusive N-acyl group migration. A monocyclic β-lactam diol undergoes Appel reaction at a primary alcohol in preference to semipinacol rearrangement. Preliminary investigations into the chemo- and stereoselective manipulation of the two carbonyl groups present in a representative 7,8-dioxo-6-azabicyclo[3.2.1]octane rearrangement product are also reported. PMID:24711140

  19. Synthetic Lethality Reveals Mechanisms of Mycobacterium tuberculosis Resistance to β-Lactams

    PubMed Central

    Lun, Shichun; Miranda, David; Kubler, Andre; Guo, Haidan; Maiga, Mariama C.; Winglee, Kathryn; Pelly, Shaaretha

    2014-01-01

    ABSTRACT Most β-lactam antibiotics are ineffective against Mycobacterium tuberculosis due to the microbe’s innate resistance. The emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains has prompted interest to repurpose this class of drugs. To identify the genetic determinants of innate β-lactam resistance, we carried out a synthetic lethality screen on a transposon mutant library for susceptibility to imipenem, a carbapenem β-lactam antibiotic. Mutations in 74 unique genes demonstrated synthetic lethality. The majority of mutations were in genes associated with cell wall biosynthesis. A second quantitative real-time PCR (qPCR)-based synthetic lethality screen of randomly selected mutants confirmed the role of cell wall biosynthesis in β-lactam resistance. The global transcriptional response of the bacterium to β-lactams was investigated, and changes in levels of expression of cell wall biosynthetic genes were identified. Finally, we validated these screens in vivo using the MT1616 transposon mutant, which lacks a functional acyl-transferase gene. Mice infected with the mutant responded to β-lactam treatment with a 100-fold decrease in bacillary lung burden over 4 weeks, while the numbers of organisms in the lungs of mice infected with wild-type bacilli proliferated. These findings reveal a road map of genes required for β-lactam resistance and validate synthetic lethality screening as a promising tool for repurposing existing classes of licensed, safe, well-characterized antimicrobials against tuberculosis. PMID:25227469

  20. A luminescent Escherichia coli biosensor for the high throughput detection of beta-lactams.

    PubMed

    Valtonen, Satu J; Kurittu, Jussi S; Karp, Matti T

    2002-04-01

    A group-specific bioluminescent Escherichia coli strain for studying the action of beta-lactam antibiotics is described. The strain contains a plasmid, pBlaLux1, in which the luciferase genes from Photorhabdus luminescens are inserted under the control of the beta-lactam-responsive element ampR/ampC from Citrobacter freundii. In the presence of beta-lactams, the bacterial cells are induced to express the luciferase enzyme and three additional enzymes generating the substrate for the luciferase reaction. This biosensor for beta-lactams does not need any substrate or cofactor additions, and the bioluminescence can be measured very sensitively in real time by using a luminometer. Basic parameters affecting the light production and induction in the gram-negative model organism E. coli SNO301/pBlaLux1 by various beta-lactams were studied. The dose-response curves were bell shaped, indicating toxic effects for the sensor strain at high concentrations of beta-lactams. Various beta-lactams had fairly different assay ranges: ampicillin, 0.05-1.0 microg/ml; piperacillin, 0.0025-25 microg/ml; imipenem, 0.0025-0.25 microg/ml; cephapirin, 0.025-2.5 microg/ml; cefoxitin, 0.0025-1.5 microg/ml; and oxacillin, 25-500 microg/ml. Also, the induction coefficients (signal over background noninduced control) varied considerably from 3 to 158 in a 2-hour assay. Different non-beta-lactam antibiotics did not cause induction. Because the assay can be automated using microplate technologies, the approach may be suitable for higher throughput analysis of beta-lactam action.

  1. Beta-lactam Antibiotics: From Antibiosis to Resistance and Bacteriology

    PubMed Central

    Kong, Kok-Fai; Schneper, Lisa; Mathee, Kalai

    2010-01-01

    SUMMARY This review focuses on the era of antibiosis that led to a better understanding of bacterial morphology, in particlar the cell wall component peptidoglycan. This is an effort to take readers on a tour de force from the concept of antibiosis, to the serepidity of antibiotics, evolution of beta-lactam development, and the molecular biology of antibiotic resistance. These areas of research have culminated in a deeper understanding of microbiology, particularly in the area of bacterial cell wall synthesis and recycling. In spite of this knowledge, which has enabled design of new even more effective therapeutics to combat bacterial infection and has provided new research tools, antibiotic resistance remains a worldwide health care problem. PMID:20041868

  2. Antimicrobial stewardship of β-lactams in intensive care units.

    PubMed

    Cotta, Menino Osbert; Roberts, Jason A; Tabah, Alexis; Lipman, Jeffrey; Vogelaers, Dirk; Blot, Stijn

    2014-05-01

    Stewardship of all antimicrobials, including β-lactam antibiotics, has gained in prominence over the last decade. Appropriate use of these agents has become vitally important; especially in the treatment and management of the critically ill. Opportunities therefore exist to develop innovations to optimise the use of antimicrobials in places like the intensive care unit. The next few years represent an important window in which routine antimicrobial stewardship principles such as surveillance of local ecology, minimising overlap of spectrum of activity and prompt de-escalation upon review of cultures can be integrated with new technologies including improved diagnostic techniques, individualised dosing strategies and computerised decision support. It is important though, that these measures to improve stewardship in the intensive care unit continue to be critically evaluated in the literature.

  3. Nitrogen metabolism in tabtoxinine-. beta. -lactam-tolerant oats

    SciTech Connect

    Knight, T.J.; Langston-Unkefer, P.J. New Mexico State Univ. Plant Genetic Engineering Lab., Las Cruces ); Sengupta-Gopalan, C. )

    1989-04-01

    Infestation of the rhizosphere of oat plants with Pseudomonas syringae pv. tabaci results in rapid death of normal oats. This is a consequence of the action of the bacterially delivered inhibitor of glutamine synthetase, tabtoxinine-{beta}-lactam (T{beta}L). Such infested plants contain no active glutamine synthetase. We have screened for a small population of oats that contain leaf glutamine synthetases that are insensitive to T{beta}L and which have increased leaf GS activity. The root GS is inactive. We have examined these plants' altered nitrogen metabolism and further characterized their novel glutamine synthetase using both biochemical and molecular biological approaches. This investigation has revealed a GS with unusual electrophoretic mobility by native PAGE.

  4. Novel reaction of elastase with cephalosporin. beta. -lactams

    SciTech Connect

    Lin, T.Y.; Williams, H.R.; Navia, M.A.; Springer, J.P.; Hoogsteen, K.; Shah, S.K.; Finke, P.E.; Doherty, J.B.; Firestone, R.A.

    1987-05-01

    Porcine pancreatic elastase (PPE) was inactivated by two cephalosporin ..beta..-lactams, 3-acetoxymethyl-7-..cap alpha..-chloro-3-cephem-4-carboxylate-1,1-dioxide t-butylester (I) and its 7-..cap alpha..-methoxy analog (II) with the first-order rate constants for inactivation, 0.023 and 0.018 s-1 respectively at pH 7.4, 25/sup 0/C. The inhibition was caused by stoichiometric binding of the compounds with PPE (KI, 80 and 30 nM at pH 7.4, respectively) followed by acylation of the active site serine with opening of the lactam ring. PPE inactivated by II (E-II) spontaneously regenerated enzyme activity with a t1/2 of 100 min at both pH 7.4 and 5.0. The reactivation of E-II was slowed with 1% SDS. The major /sup 14/C-labeled tryptic peptide of PPE modified with (/sup 14/C)MeO-labeled II had the amino acid composition of the sequence Ser182 to Arg211. PPE inactivation with I did not reactivate but showed a time-dependent resistance to reactivation by treatment with 0.5 M NH2OH at pH 7.5 and 37/sup 0/C for 10 min. The acid hydrolyzate of PPE-I contained 5 residues of histidine/mole rather than 6 for native PPE. PPE crystals soaked with I in 35% PEG 4000, 0.1 M NaOAc, pH 5.0 were subjected to high resolution x-ray diffraction analysis. The cross-linking of the active site at Ser188 OH and His45 N/sup 2/ by a 2-substituted, 5-methylene-1,3-thiazine dioxide was clearly demonstrated.

  5. Practical Electrochemical Anodic Oxidation of Polycyclic Lactams for Late Stage Functionalization**

    PubMed Central

    Frankowski, Kevin J.; Liu, Ruzhang; Milligan, Gregory L.; Moeller, Kevin D.

    2015-01-01

    Electrochemistry provides a powerful tool for the late-stage functionalization of complex lactams. A two-stage protocol for converting lactams, many of which are preparable through the intramolecular Schmidt reaction of keto azides, is presented. In the first step, anodic oxidation in MeOH using a repurposed power source provides a convenient route to lactams bearing a methoxy group adjacent to nitrogen. Treatment of these intermediates with a Lewis acid in DCM permits the regeneration of a reactive acyliminium ion that is then reacted with a range of nucleophilic species. PMID:26371961

  6. Is the addition of aminoglycosides to beta-lactams in cancer patients with febrile neutropenia needed?

    PubMed

    Contreras, Valeria; Sepúlveda, Sebastián; Heredia, Ana

    2016-02-24

    It is still controversial if the combined use of beta-lactam antibiotics and aminoglycosides has advantages over broad-spectrum beta-lactam monotherapy for the empirical treatment of cancer patients with febrile neutropenia. Searching in Epistemonikos database, which is maintained by screening 30 databases, we identified three systematic reviews including 14 pertinent randomized trials. We combined the evidence using meta-analysis and generated a summary of findings table following the GRADE approach. We concluded the combination of beta-lactam antibiotics and aminoglycosides probably does not lead to a reduced mortality in febrile neutropenic cancer patients and it might increase nephrotoxicity.

  7. Is the addition of aminoglycosides to beta-lactams in cancer patients with febrile neutropenia needed?

    PubMed

    Contreras, Valeria; Sepúlveda, Sebastián; Heredia, Ana

    2016-01-01

    It is still controversial if the combined use of beta-lactam antibiotics and aminoglycosides has advantages over broad-spectrum beta-lactam monotherapy for the empirical treatment of cancer patients with febrile neutropenia. Searching in Epistemonikos database, which is maintained by screening 30 databases, we identified three systematic reviews including 14 pertinent randomized trials. We combined the evidence using meta-analysis and generated a summary of findings table following the GRADE approach. We concluded the combination of beta-lactam antibiotics and aminoglycosides probably does not lead to a reduced mortality in febrile neutropenic cancer patients and it might increase nephrotoxicity. PMID:26938974

  8. Synthesis of novel N-cyclopentenyl-lactams using the Aubé reaction

    PubMed Central

    Shinde, Madhuri V; Ople, Rohini S; Sangtani, Ekta; Gonnade, Rajesh

    2015-01-01

    Summary A novel and convenient method utilizing the Aubé reaction to access a new class of compounds that are similar to carbocyclic nucleosides is reported. The azido alcohol derived from Vince lactam undergoes the Aubé reaction with various cyclic ketones to give cyclopentenyl-substituted lactams. Upon dihydroxylation, this affords the N-cyclopentenyl-lactam compounds in racemic form. Given the numerous uses of nucleosides and related compounds, we were interested in the synthesis of carbocylic nucleoside mimics. The attempts and results are described herein. PMID:26199661

  9. Synthesis of a tricyclic lactam via Beckmann rearrangement and ring-rearrangement metathesis as key steps

    PubMed Central

    Ravikumar, Ongolu; Majhi, Jadab

    2015-01-01

    Summary A tricyclic lactam is reported in a four step synthesis sequence via Beckmann rearrangement and ring-rearrangement metathesis as key steps. Here, we used a simple starting material such as dicyclopentadiene. PMID:26425207

  10. Molecular Targets of β-Lactam-Based Antimicrobials: Beyond the Usual Suspects

    PubMed Central

    Konaklieva, Monika I.

    2014-01-01

    The common practice in antibacterial drug development has been to rapidly make an attempt to find ever-more stable and broad-spectrum variants for a particular antibiotic, once a drug resistance for that antibiotic is detected. We are now facing bacterial resistance toward our clinically relevant antibiotics of such a magnitude that the conversation for antimicrobial drug development ought to include effective new antibiotics with alternative mechanisms of action. The electrophilic β-lactam ring is amenable for the inhibition of different enzyme classes by a suitable decoration of the core scaffold. Monocyclic β-lactams lacking an ionizable group at the lactam nitrogen exhibit target preferences toward bacterial enzymes important for resistance and virulence. The present review intends to draw attention to the versatility of the β-lactams as antimicrobials with “unusual” molecular targets. PMID:27025739

  11. Subtleties in practical application of prolonged infusion of β-lactam antibiotics.

    PubMed

    De Waele, Jan J; Lipman, Jeffrey; Carlier, Mieke; Roberts, Jason A

    2015-05-01

    Prolonged infusion (PI) of β-lactam antibiotics is increasingly used in order to optimise antibiotic exposure in critically ill patients. Physicians are often not aware of a number of subtleties that may jeopardise the treatment. In this clinically based paper, we stress pragmatic issues, such as the importance of a loading dose before PI, and discuss a number of important practicalities that are mandatory to benefit from the pharmacokinetic advantages of prolonged β-lactam antibiotic administration.

  12. Chemical and microbiologic aspects of penems, a distinct class of beta-lactams: focus on faropenem.

    PubMed

    Hamilton-Miller, Jeremy M T

    2003-11-01

    Many beta-lactam antimicrobials were developed between the 1960s and 1980s, with continuing development driven by the emergence of microbial resistance. Penems form a discrete class of beta-lactams that comprises structural hybrids of penicillins (penams) and cephalosporins (cephems). The chemistry and microbiology of the representative penems MEN 10700, ritipenem, CGP 31608, sulopenem, BRL 42715, and faropenem are reviewed. Particular emphasis is placed on faropenem, which is in late clinical development. PMID:14620395

  13. Aureoverticillactam, a novel 22-atom macrocyclic lactam from the marine actinomycete Streptomyces aureoverticillatus.

    PubMed

    Mitchell, Scott S; Nicholson, Benjamin; Teisan, Sy; Lam, Kin S; Potts, Barbara C M

    2004-08-01

    During the course of our screening program designed to discover novel anticancer and anti-infective agents from marine microorganisms, a strain of Streptomyces aureoverticillatus (NPS001583) isolated from a marine sediment was found to produce a novel macrocyclic lactam with cytotoxicity against various tumor cell lines. Using extensive MS, UV, and NMR spectral analyses, the structure has been established as compound 1, aureoverticillactam, a 22-atom macrocyclic lactam incorporating both triene and tetraene conjugated olefins. PMID:15332863

  14. Stereoselective Synthesis of Ezetimibe via Cross-Metathesis of Homoallylalcohols and α-Methylidene-β-Lactams.

    PubMed

    Humpl, Marek; Tauchman, Jiří; Topolovčan, Nikola; Kretschmer, Jan; Hessler, Filip; Císařová, Ivana; Kotora, Martin; Veselý, Jan

    2016-09-01

    Ru-catalyzed cross-metathesis (CM) reaction between β-arylated α-methylidene-β-lactams and terminal olefins was developed. The CM reaction is effectively catalyzed with Hoveyda-Grubbs second-generation catalyst affording corresponding α-alkylidene-β-aryl-β-lactams in good isolated yields (41-83%) with exclusive Z-selectivity. The developed protocol was successfully applied for stereoselective preparation of Ezetimibe, the commercial cholesterol absorption inhibitor. PMID:27494518

  15. Selective trihydroxyazepane NagZ inhibitors increase sensitivity of Pseudomonas aeruginosa to β-lactams.

    PubMed

    Mondon, Martine; Hur, Soo; Vadlamani, Grishma; Rodrigues, Prerana; Tsybina, Polina; Oliver, Antonio; Mark, Brian L; Vocadlo, David J; Blériot, Yves

    2013-12-01

    AmpC β-lactamase confers resistance to β-lactam antibiotics in many Gram negative bacteria. Inducible expression of AmpC requires an N-acetylglucosaminidase termed NagZ. Here we describe the synthesis and characterization of hydroxyazepane inhibitors of NagZ. We find that these inhibitors enhance the susceptibility of clinically relevant Pseudomonas aeruginosa to β-lactams. PMID:24136176

  16. Synthesis and in vitro anti-tubercular evaluation of 1,2,3-triazole tethered β-lactam-ferrocene and β-lactam-ferrocenylchalcone chimeric scaffolds.

    PubMed

    Kumar, Kewal; Singh, Pardeep; Kremer, Laurent; Guérardel, Yann; Biot, Christophe; Kumar, Vipan

    2012-05-21

    Twenty different triazoles were prepared to probe the anti-tubercular structure-activity relationships (SAR) within the β-lactam-ferrocene-triazole conjugate family. The compounds have been synthesized by copper-catalyzed "click chemistry". In vitro anti-tubercular activity was determined for each compound but the synthesized hybrids failed to inhibit Mycobacterium tuberculosis growth even at high doses. The manuscript assumes significance as this is the first report on the inclusion of ferrocene nucleus in the well established β-lactam family via triazole linkers with reputed physicochemical profiles.

  17. Mass Spectral Profile for Rapid Differentiating Beta-Lactams from Their Ring-Opened Impurities

    PubMed Central

    Wang, Hecheng; Huang, Haiwei; Cao, Jin; Chui, Dehua

    2015-01-01

    High performance liquid chromatography tandem mass spectrometry (HPLC MS) has been widely used for β-lactam antibiotics determination. However, its application to identify impurities of these frequently used drugs is not sufficient at present. In this job, characteristic profiles of the collision induced dissociation (CID) spectra of both β-lactams and ring-opened β-lactams were extracted from the MS data of six β-lactam antibiotics and their forty-five impurities, and were confirmed by the MS data reported in the literature. These characteristics have been successfully applied to rapid differentiation of β-lactam and ring-opened β-lactam impurities in cefixime, cefdinir, and cefaclor. However, these characteristic profiles can only be obtained under low activating voltage. They did not display in the high energy activated CID spectra. Diagnostic fragmentations for determining the localization of double bond and substituents on the thiazine ring and the side chain were also observed. In addition, several characteristic fragmentations are hopeful to be used to differentiate the configurations of C-2 on the thiazine ring of ring-opened impurities, which is generally disadvantageous of mass spectrometry. Taken together, forty-five impurities were identified from the capsules of cefixime, cefdinir, and cefaclor. PMID:26090434

  18. Novel Aspects on the Preparation of Spirocyclic and Fused Unusual β-Lactams

    NASA Astrophysics Data System (ADS)

    Alcaide, Benito; Almendros, Pedro

    β-Lactam antibiotics have occupied a central role in the fight against pathogenic bacteria and the subsequent rise in quality of life for the world population as a whole. However, the extensive use of common β-lactam antibiotics such as penicillins and cephalosporins in medicine has resulted in an increasing number of resistant strains of bacteria through mutation and β-lactamase gene transfer. The resistance of bacteria to the classical β-lactam antibiotics can be overcome, e.g., by using novel β-lactam moieties in drugs, which show much higher stability towards these resistance bacteria. In addition, there are many important nonantibiotic uses of 2-azetidinones in fields ranging from enzyme inhibition to gene activation. These biological activities, combined with the use of these products as starting materials to prepare α- and β-amino acids, alkaloids, heterocycles, taxoids, and other types of compounds of biological and medicinal interest, provide the motivation to explore new methodologies for the synthesis of substances based on the β-lactam core. The aim of this chapter is to provide a survey of the types of reactions used to prepare nonconventional spirocyclic and fused β-lactams, concentrating on the advances that have been made in the last decade, particularly in the last 5 years. We will draw special attention to radical cyclizations, cycloaddition reactions, and transition metal-catalyzed reactions.

  19. Mass Spectral Profile for Rapid Differentiating Beta-Lactams from Their Ring-Opened Impurities.

    PubMed

    Wang, Hecheng; Huang, Haiwei; Cao, Jin; Chui, Dehua; Xiao, Shengyuan

    2015-01-01

    High performance liquid chromatography tandem mass spectrometry (HPLC MS) has been widely used for β-lactam antibiotics determination. However, its application to identify impurities of these frequently used drugs is not sufficient at present. In this job, characteristic profiles of the collision induced dissociation (CID) spectra of both β-lactams and ring-opened β-lactams were extracted from the MS data of six β-lactam antibiotics and their forty-five impurities, and were confirmed by the MS data reported in the literature. These characteristics have been successfully applied to rapid differentiation of β-lactam and ring-opened β-lactam impurities in cefixime, cefdinir, and cefaclor. However, these characteristic profiles can only be obtained under low activating voltage. They did not display in the high energy activated CID spectra. Diagnostic fragmentations for determining the localization of double bond and substituents on the thiazine ring and the side chain were also observed. In addition, several characteristic fragmentations are hopeful to be used to differentiate the configurations of C-2 on the thiazine ring of ring-opened impurities, which is generally disadvantageous of mass spectrometry. Taken together, forty-five impurities were identified from the capsules of cefixime, cefdinir, and cefaclor. PMID:26090434

  20. Mass Spectral Profile for Rapid Differentiating Beta-Lactams from Their Ring-Opened Impurities.

    PubMed

    Wang, Hecheng; Huang, Haiwei; Cao, Jin; Chui, Dehua; Xiao, Shengyuan

    2015-01-01

    High performance liquid chromatography tandem mass spectrometry (HPLC MS) has been widely used for β-lactam antibiotics determination. However, its application to identify impurities of these frequently used drugs is not sufficient at present. In this job, characteristic profiles of the collision induced dissociation (CID) spectra of both β-lactams and ring-opened β-lactams were extracted from the MS data of six β-lactam antibiotics and their forty-five impurities, and were confirmed by the MS data reported in the literature. These characteristics have been successfully applied to rapid differentiation of β-lactam and ring-opened β-lactam impurities in cefixime, cefdinir, and cefaclor. However, these characteristic profiles can only be obtained under low activating voltage. They did not display in the high energy activated CID spectra. Diagnostic fragmentations for determining the localization of double bond and substituents on the thiazine ring and the side chain were also observed. In addition, several characteristic fragmentations are hopeful to be used to differentiate the configurations of C-2 on the thiazine ring of ring-opened impurities, which is generally disadvantageous of mass spectrometry. Taken together, forty-five impurities were identified from the capsules of cefixime, cefdinir, and cefaclor.

  1. One ring to rule them all: Current trends in combating bacterial resistance to the β-lactams.

    PubMed

    King, Dustin T; Sobhanifar, Solmaz; Strynadka, Natalie C J

    2016-04-01

    From humble beginnings of a contaminated petri dish, β-lactam antibiotics have distinguished themselves among some of the most powerful drugs in human history. The devastating effects of antibiotic resistance have nevertheless led to an "arms race" with disquieting prospects. The emergence of multidrug resistant bacteria threatens an ever-dwindling antibiotic arsenal, calling for new discovery, rediscovery, and innovation in β-lactam research. Here the current state of β-lactam antibiotics from a structural perspective was reviewed. PMID:26813250

  2. Alpha-melanocyte stimulating hormone inhibits monocytes adhesion to vascular endothelium

    PubMed Central

    Yang, Yang; Zhang, Weihua; Meng, Lin; Yu, Haitao; Lu, Na; Fu, Gang

    2015-01-01

    Inflammation and its subsequent endothelial dysfunction have been reported to play a pivotal role in the initiation and progression of chronic vascular diseases. Inhibiting the attachment of monocytes to endothelium is a potential therapeutic strategy for vascular diseases treatment. α-Melanocyte stimulating hormone is generated from a precursor hormone called proopiomelanocortin by post-translational processing. However, whether α-melanocyte stimulating hormone plays a role in regulating endothelial inflammation is still unknown. In this study, the effects of α-melanocyte stimulating hormone on endothelial inflammation in human umbilical vein endothelial cell lines were investigated. And the result indicated that α-melanocyte stimulating hormone inhibits the expression of endothelial adhesion molecules, including vascular adhesion molecule-1 and E-selectin, thereby attenuating the adhesion of THP-1 cells to the surface of endothelial cells. Mechanistically, α-melanocyte stimulating hormone was found to inhibit NF-κB transcriptional activity. Finally, we found that the effect of α-melanocyte stimulating hormone on endothelial inflammation is dependent on its receptor melanocortin receptor 1. PMID:25898835

  3. Tyrosol and Its Analogues Inhibit Alpha-Melanocyte-Stimulating Hormone Induced Melanogenesis

    PubMed Central

    Wen, Kuo-Ching; Chang, Chih-Shiang; Chien, Yin-Chih; Wang, Hsiao-Wen; Wu, Wan-Chen; Wu, Chin-Sheng; Chiang, Hsiu-Mei

    2013-01-01

    Melanin is responsible for skin color and plays a major role in defending against harmful external factors such as ultraviolet (UV) irradiation. Tyrosinase is responsible for the critical steps of melanogenesis, including the rate-limiting step of tyrosine hydroxylation. The mechanisms of action of skin hypopigmenting agents are thought to be based on the ability of a given agent to inhibit the activity of tyrosinase and, hence, down regulate melanin synthesis. Tyrosol and its glycoside, salidroside, are active components of Rhodiola rosea, and in our preliminary study we found that Rhodiola rosea extract inhibited melanogenesis. In this study, we examined the effects of tyrosol and its analogues on melanin synthesis. We found that treatment of B16F0 cells to tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), 2-hydroxyphenylacetic acid (7), or salidroside (11) resulted in a reduction in melanin content and inhibition of tyrosinase activity as well as its expression. Tyrosol (1), 4-hydroxyphenylacetic acid (5) and 2-hydroxyphenylacetic acid (7) suppressed MC1R expression. Tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), and 2-hydroxyphenylacetic acid (7) inhibited α-MSH induced TRP-1 expression, but salidroside (11) did not. All the compounds did not affect MITF and TRP-2 expression. Furthermore, we found that the cell viability of tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), and 2-hydroxyphenylacetic acid (7) at concentrations below 4 mM and salidroside (11) at concentrations below 0.5 mM were higher than 90%. The compounds exhibited metal-coordinating interactions with copper ion in molecular docking with tyrosinase. Our results suggest that tyrosol, 4-hydroxyphenylacetic acid, 3-hydroxyphenylacetic acid, 2-hydroxyphenylacetic acid, and salidroside are potential hypopigmenting agents. PMID:24287915

  4. Tyrosol and its analogues inhibit alpha-melanocyte-stimulating hormone induced melanogenesis.

    PubMed

    Wen, Kuo-Ching; Chang, Chih-Shiang; Chien, Yin-Chih; Wang, Hsiao-Wen; Wu, Wan-Chen; Wu, Chin-Sheng; Chiang, Hsiu-Mei

    2013-01-01

    Melanin is responsible for skin color and plays a major role in defending against harmful external factors such as ultraviolet (UV) irradiation. Tyrosinase is responsible for the critical steps of melanogenesis, including the rate-limiting step of tyrosine hydroxylation. The mechanisms of action of skin hypopigmenting agents are thought to be based on the ability of a given agent to inhibit the activity of tyrosinase and, hence, down regulate melanin synthesis. Tyrosol and its glycoside, salidroside, are active components of Rhodiola rosea, and in our preliminary study we found that Rhodiola rosea extract inhibited melanogenesis. In this study, we examined the effects of tyrosol and its analogues on melanin synthesis. We found that treatment of B16F0 cells to tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), 2-hydroxyphenylacetic acid (7), or salidroside (11) resulted in a reduction in melanin content and inhibition of tyrosinase activity as well as its expression. Tyrosol (1), 4-hydroxyphenylacetic acid (5) and 2-hydroxyphenylacetic acid (7) suppressed MC1R expression. Tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), and 2-hydroxyphenylacetic acid (7) inhibited α-MSH induced TRP-1 expression, but salidroside (11) did not. All the compounds did not affect MITF and TRP-2 expression. Furthermore, we found that the cell viability of tyrosol (1), 4-hydroxyphenylacetic acid (5), 3-hydroxyphenylacetic acid (6), and 2-hydroxyphenylacetic acid (7) at concentrations below 4 mM and salidroside (11) at concentrations below 0.5 mM were higher than 90%. The compounds exhibited metal-coordinating interactions with copper ion in molecular docking with tyrosinase. Our results suggest that tyrosol, 4-hydroxyphenylacetic acid, 3-hydroxyphenylacetic acid, 2-hydroxyphenylacetic acid, and salidroside are potential hypopigmenting agents. PMID:24287915

  5. The bactericidal activity of β-lactam antibiotics is increased by metabolizable sugar species.

    PubMed

    Thorsing, Mette; Bentin, Thomas; Givskov, Michael; Tolker-Nielsen, Tim; Goltermann, Lise

    2015-10-01

    Here, the influence of metabolizable sugars on the susceptibility of Escherichia coli to β-lactam antibiotics was investigated. Notably, monitoring growth and survival of mono- and combination-treated planktonic cultures showed a 1000- to 10 000-fold higher antibacterial efficacy of carbenicillin and cefuroxime in the presence of certain sugars, whereas other metabolites had no effect on β-lactam sensitivity. This effect was unrelated to changes in growth rate. Light microscopy and flow cytometry profiling revealed that bacterial filaments, formed due to β-lactam-mediated inhibition of cell division, rapidly appeared upon β-lactam mono-treatment and remained stable for up to 18 h. The presence of metabolizable sugars in the medium did not change the rate of filamentation, but led to lysis of the filaments within a few hours. No lysis occurred in E. coli mutants unable to metabolize the sugars, thus establishing sugar metabolism as an important factor influencing the bactericidal outcome of β-lactam treatment. Interestingly, the effect of sugar on β-lactam susceptibility was suppressed in a strain unable to synthesize the nutrient stress alarmone (p)ppGpp. Here, to the best of our knowledge, we demonstrate for the first time a specific and significant increase in β-lactam sensitivity due to sugar metabolism in planktonic, exponentially growing bacteria, unrelated to general nutrient availability or growth rate. Understanding the mechanisms underlying the nutritional influences on antibiotic sensitivity is likely to reveal new proteins or pathways that can be targeted by novel compounds, adding to the list of pharmacodynamic adjuvants that increase the efficiency and lifespan of conventional antibiotics.

  6. Energetic, Structural, and Antimicrobial Analyses of [beta]-Lactam Side Chain Recognition by [beta]-Lactamases

    SciTech Connect

    Caselli, E.; Powers, R.A.; Blaszczak, L.C.; Wu, C.Y.E.; Prati, F.; Shoichet, B.K.

    2010-03-05

    Penicillins and cephalosporins are among the most widely used and successful antibiotics. The emergence of resistance to these {beta}-lactams, most often through bacterial expression of {beta}-lactamases, threatens public health. To understand how {beta}-lactamases recognize their substrates, it would be helpful to know their binding energies. Unfortunately, these have been difficult to measure because {beta}-lactams form covalent adducts with {beta}-lactamases. This has complicated functional analyses and inhibitor design. To investigate the contribution to interaction energy of the key amide (R1) side chain of {beta}-lactam antibiotics, eight acylglycineboronic acids that bear the side chains of characteristic penicillins and cephalosporins, as well as four other analogs, were synthesized. These transition-state analogs form reversible adducts with serine {beta}-lactamases. Therefore, binding energies can be calculated directly from K{sub i} values. The K{sub i} values measured span four orders of magnitude against the Group I {beta}-lactamase AmpC and three orders of magnitude against the Group II {beta}-lactamase TEM-1. The acylglycineboronic acids have K{sub i} values as low as 20 nM against AmpC and as low as 390 nM against TEM-1. The inhibitors showed little activity against serine proteases, such as chymotrypsin. R1 side chains characteristic of {beta}-lactam inhibitors did not have better affinity for AmpC than did side chains characteristic of {beta}-lactam substrates. Two of the inhibitors reversed the resistance of pathogenic bacteria to {beta}-lactams in cell culture. Structures of two inhibitors in their complexes with AmpC were determined by X-ray crystallography to 1.90 {angstrom} and 1.75 {angstrom} resolution; these structures suggest interactions that are important to the affinity of the inhibitors. Acylglycineboronic acids allow us to begin to dissect interaction energies between {beta}-lactam side chains and {beta}-lactamases. Surprisingly

  7. Heat inactivation of beta-lactam antibiotics in milk.

    PubMed

    Zorraquino, M A; Roca, M; Fernandez, N; Molina, M P; Althaus, R

    2008-06-01

    The presence of residues of antimicrobial substances in milk is one of the main concerns of the milk industry, as it poses a risk of toxicity to public health, and can seriously influence the technological properties of milk and dairy products. Moreover, the information available on the thermostability characteristics of these residues, particularly regarding the heat treatments used in control laboratories and the dairy industry, is very scarce. The aim of the study was, therefore, to analyze the effect of different heat treatments (40 degrees C for 10 min, 60 degrees C for 30 min, 83 degrees C for 10 min, 120 degrees C for 20 min, and 140 degrees C for 10 s) on milk samples fortified with three concentrations of nine beta-lactam antibiotics (penicillin G: 3, 6, and 12 microg/liter; ampicillin: 4, 8, and 16 microg/liter; amoxicillin: 4, 8, and 16 microg/liter; cloxacillin: 60, 120, and 240 microg/liter; cefoperazone: 55, 110, and 220 microg/liter; cefquinome: 100, 200, and 400 microg/liter; cefuroxime: 65, 130, and 260 microg/liter; cephalexin: 80, 160, and 220 microg/ liter; and cephalonium: 15, 30, and 60 microg/liter). The method used was a bioassay based on the inhibition of Geobacillus stearothermophilus var. calidolactis. The results showed that heating milk samples at 40 degrees C for 10 min hardly produced any heat inactivation at all, while the treatment at 83 degrees C for 10 min caused a 20% loss in penicillin G, 27% in cephalexin, and 35% in cefuroxime. Of the three dairy industry heat treatments studied in this work, low pasteurization (60 degrees C for 30 min) and treatment at 140 degrees C for 10 s only caused a small loss of antimicrobial activity, whereas classic sterilization (120 degrees C for 20 min) showed a high level of heat inactivation of over 65% for penicillins and 90% for cephalosporins. PMID:18592745

  8. Heat inactivation of beta-lactam antibiotics in milk.

    PubMed

    Zorraquino, M A; Roca, M; Fernandez, N; Molina, M P; Althaus, R

    2008-06-01

    The presence of residues of antimicrobial substances in milk is one of the main concerns of the milk industry, as it poses a risk of toxicity to public health, and can seriously influence the technological properties of milk and dairy products. Moreover, the information available on the thermostability characteristics of these residues, particularly regarding the heat treatments used in control laboratories and the dairy industry, is very scarce. The aim of the study was, therefore, to analyze the effect of different heat treatments (40 degrees C for 10 min, 60 degrees C for 30 min, 83 degrees C for 10 min, 120 degrees C for 20 min, and 140 degrees C for 10 s) on milk samples fortified with three concentrations of nine beta-lactam antibiotics (penicillin G: 3, 6, and 12 microg/liter; ampicillin: 4, 8, and 16 microg/liter; amoxicillin: 4, 8, and 16 microg/liter; cloxacillin: 60, 120, and 240 microg/liter; cefoperazone: 55, 110, and 220 microg/liter; cefquinome: 100, 200, and 400 microg/liter; cefuroxime: 65, 130, and 260 microg/liter; cephalexin: 80, 160, and 220 microg/ liter; and cephalonium: 15, 30, and 60 microg/liter). The method used was a bioassay based on the inhibition of Geobacillus stearothermophilus var. calidolactis. The results showed that heating milk samples at 40 degrees C for 10 min hardly produced any heat inactivation at all, while the treatment at 83 degrees C for 10 min caused a 20% loss in penicillin G, 27% in cephalexin, and 35% in cefuroxime. Of the three dairy industry heat treatments studied in this work, low pasteurization (60 degrees C for 30 min) and treatment at 140 degrees C for 10 s only caused a small loss of antimicrobial activity, whereas classic sterilization (120 degrees C for 20 min) showed a high level of heat inactivation of over 65% for penicillins and 90% for cephalosporins.

  9. Alkyl Gallates, Intensifiers of β-Lactam Susceptibility in Methicillin-Resistant Staphylococcus aureus

    PubMed Central

    Shibata, Hirofumi; Kondo, Kyoko; Katsuyama, Ryo; Kawazoe, Kazuyoshi; Sato, Yoichi; Murakami, Kotaro; Takaishi, Yoshihisa; Arakaki, Naokatu; Higuti, Tomihiko

    2005-01-01

    We found that ethyl gallate purified from a dried pod of tara (Caesalpinia spinosa) intensified β-lactam susceptibility in methicillin-resistant and methicillin-sensitive strains of Staphylococcus aureus (MRSA and MSSA strains, respectively). This compound and several known alkyl gallates were tested with MRSA and MSSA strains to gain new insights into their structural functions in relation to antimicrobial and β-lactam susceptibility-intensifying activities. The maximum activity of alkyl gallates against MRSA and MSSA strains occurred at 1-nonyl and 1-decyl gallate, with an MIC at which 90% of the isolates tested were inhibited of 15.6 μg/ml. At concentrations lower than the MIC, alkyl gallates synergistically elevated the susceptibility of MRSA and MSSA strains to β-lactam antibiotics. Such a synergistic activity of the alkyl gallates appears to be specific for β-lactam antibiotics, because no significant changes were observed in the MICs of other classes of antibiotics examined in this study. The length of the alkyl chain was also associated with the modifying activity of the alkyl gallates, and the optimum length was C5 to C6. The present work clearly demonstrates that the length of the alkyl chain has a key role in the elevation of susceptibility to β-lactam antibiotics. PMID:15673731

  10. Synthesis of ¹⁸F-labelled β-lactams by using the Kinugasa reaction.

    PubMed

    Zlatopolskiy, Boris D; Krapf, Philipp; Richarz, Raphael; Frauendorf, Holm; Mottaghy, Felix M; Neumaier, Bernd

    2014-04-14

    Owing to their broad spectrum of biological activities and low toxicity, β-lactams are attractive lead structures for the design of novel molecular probes. However, the synthesis of positron emission tomography (PET)-isotope-labelled β-lactams has not yet been reported. Herein, we describe the simple preparation of radiofluorinated β-lactams by using the fast Kinugasa reaction between (18)F-labelled nitrone [(18)F]-1 and alkynes of different reactivity. Additionally, (18)F-labelled fused β-lactams were obtained through the reaction of a cyclic nitrone 7 with radiofluorinated alkynes [(18)F]-6 a,b. Radiochemical yields of the Kinugasa reaction products could be significantly increased by the use of different Cu(I) ligands, which additionally allowed a reduction in the amount of precursor and/or reaction time. Model radiofluorinated β-lactam-peptide and protein conjugates ([(18)F]-10 and (18)F-labelled BSA conjugate) were efficiently obtained in high yield under mild conditions (aq. MeCN, ambient temperature) within a short reaction time, demonstrating the suitability of the developed method for radiolabelling of sensitive molecules such as biopolymers.

  11. A gold immunochromatographic assay for the rapid and simultaneous detection of fifteen β-lactams

    NASA Astrophysics Data System (ADS)

    Chen, Yanni; Wang, Yongwei; Liu, Liqiang; Wu, Xiaoling; Xu, Liguang; Kuang, Hua; Li, Aike; Xu, Chuanlai

    2015-10-01

    A novel gold immunochromatographic assay (GICA) based on anti-β-lactam receptors was innovatively developed that successfully allowed rapid and simultaneous detection of fifteen β-lactams in milk samples in 5-10 minutes. By replacing the antibodies used in traditional GICA with anti-β-lactam receptors, the difficulty in producing broad specific antibodies against β-lactams was overcome. Conjugates of ampicillin with BSA and goat anti-mouse immunoglobulin (IgG) were immobilized onto the test and control lines on the nitrocellulose membrane, respectively. Since goat anti-mouse IgG does not combine with receptors, negative serum from mice labelled with gold nanoparticles (GNP) was mixed with GNP-labelled receptors. Results were obtained within 20 min using a paper-based sensor. The utility of the assay was confirmed by the analysis of milk samples. The limits of detection (LOD) for amoxicillin, ampicillin, penicillin G, penicillin V, cloxacillin, dicloxacillin, nafcillin, oxacillin, cefaclor, ceftezole, cefotaxime, ceftiofur, cefoperazone, cefathiamidine, and cefepime were 0.25, 0.5, 0.5, 0.5, 1, 5, 5, 10, 25, 10, 100, 10, 5, 5, and 2 ng mL-1, respectively, which satisfies the maximum residue limits (MRL) set by the European Union (EU). In conclusion, our newly developed GICA-based anti-β-lactam receptor assay provides a rapid and effective method for one-site detection of multiple β-lactams in milk samples.A novel gold immunochromatographic assay (GICA) based on anti-β-lactam receptors was innovatively developed that successfully allowed rapid and simultaneous detection of fifteen β-lactams in milk samples in 5-10 minutes. By replacing the antibodies used in traditional GICA with anti-β-lactam receptors, the difficulty in producing broad specific antibodies against β-lactams was overcome. Conjugates of ampicillin with BSA and goat anti-mouse immunoglobulin (IgG) were immobilized onto the test and control lines on the nitrocellulose membrane, respectively

  12. Lactam based 7-amino suberoylamide hydroxamic acids as potent HDAC inhibitors.

    PubMed

    Taddei, Maurizio; Cini, Elena; Giannotti, Luca; Giannini, Giuseppe; Battistuzzi, Gianfranco; Vignola, Davide; Vesci, Loredana; Cabri, Walter

    2014-01-01

    A series of SAHA-like molecules were prepared introducing different lactam-carboxyamides in position 7 of the suberoylanilide skeleton. The activity against different HDAC isoforms was tested and the data compared with the corresponding linear products, without substituent in position 7. In general, this modification provided an effective reinforcement of in vitro activity. While the lactam size or the CO/NH group orientation did not strongly influence the inhibition, the contemporary modification of the suberoylamide fragment gave vary active variants in the lactam series, with compound 28 (ST8078AA1) that showed IC50 values between 2 and 10nM against all Class I HDAC isoforms, demonstrating it to be a large spectrum pan-inhibitor. This strong affinity with HDAC was also confirmed by the value of IC50=0.5μM against H460 cells, ranking 28 as one of the most potent HDAC inhibitors described so far. PMID:24345446

  13. Isosulfazecin, a new beta-lactam antibiotic, produced by an acidophilic pseudomonad. Fermentation, isolation and characterization.

    PubMed

    Kintaka, K; Haibara, K; Asai, M; Imada, A

    1981-09-01

    A novel beta-lactam antibiotic, isosulfazecin (iSZ), was found to be produced by an acidophilic pseudomonad, Pseudomonas mesoacidophila sp. nov. iSZ was produced in parallel with bacterial growth in nutrient broth containing glycerol and sodium thiosulfate under aerated conditions. iSZ was isolated by chromatography on activated charcoal and anion-exchangers and crystallized from 70% aqueous methanol. The molecular formula was determined to be C12H20N4O9S from physiochemical data. The IR and NMR spectra suggested that iSZ has a beta-lactam ring, methoxyl and sulfonate groups. On acid hydrolysis, it gave L-alanine and D-glutamic acid. iSZ is an epimeric isomer of sulfazecin. iSZ was weakly active against Gram-positive and -negative bacteria, and was strongly active against mutants hypersensitive to beta-lactam antibiotics. PMID:7328050

  14. Enantioselective construction of quaternary N-heterocycles by palladium-catalysed decarboxylative allylic alkylation of lactams

    NASA Astrophysics Data System (ADS)

    Behenna, Douglas C.; Liu, Yiyang; Yurino, Taiga; Kim, Jimin; White, David E.; Virgil, Scott C.; Stoltz, Brian M.

    2012-02-01

    The enantioselective synthesis of nitrogen-containing heterocycles (N-heterocycles) represents a substantial chemical research effort and resonates across numerous disciplines, including the total synthesis of natural products and medicinal chemistry. In this Article, we describe the highly enantioselective palladium-catalysed decarboxylative allylic alkylation of readily available lactams to form 3,3-disubstituted pyrrolidinones, piperidinones, caprolactams and structurally related lactams. Given the prevalence of quaternary N-heterocycles in biologically active alkaloids and pharmaceutical agents, we envisage that our method will provide a synthetic entry into the de novo asymmetric synthesis of such structures. As an entry for these investigations we demonstrate how the described catalysis affords enantiopure quaternary lactams that intercept synthetic intermediates previously used in the synthesis of the Aspidosperma alkaloids quebrachamine and rhazinilam, but that were previously only available by chiral auxiliary approaches or as racemic mixtures.

  15. Redefining the Role of psr in β-Lactam Resistance and Cell Autolysis of Enterococcus hirae

    PubMed Central

    Sapunaric, Frédéric; Franssen, Christine; Stefanic, Patrick; Amoroso, Ana; Dardenne, Olivier; Coyette, Jacques

    2003-01-01

    The contribution of penicillin-binding protein 5 (PBP5) and the PBP5 synthesis repressor (Psr) to the β-lactam resistance, growth, and cell autolysis of wild-type strain ATCC 9790 and resistant strain R40 of Enterococcus hirae was investigated by disruption or substitution of the corresponding pbp5 and psr genes by Campbell-type recombination. The resulting modifications were confirmed by hybridization and PCR. The low susceptibility of E. hirae to β-lactams was confirmed to be largely dependent on the presence of PBP5. However, against all expectations, inactivation of psr in ATCC 9790 or complementation of R40 cells with psr did not modify the susceptibility to benzylpenicillin or the growth and cell autolysis rates. These results indicated that the psr gene does not seem to be involved in the regulation of PBP5 synthesis and consequently in β-lactam resistance or in the regulation of cell autolysis in E. hirae. PMID:14526002

  16. Resistance patterns to beta-lactams and quinolones in clinical isolates of bacteria from Cuban hospitals.

    PubMed

    Gonzáles, I; Niebla, A; Vallin, C

    1995-01-01

    The resistance patterns to 26 beta-lactams and 8 quinolones of clinical isolates from Cuban hospitals were evaluated using the disk susceptibility test, according to the NCCLS guidelines (1992). The genera studied were Escherichia sp (320), Enterobacter sp (10), Klebsiella sp (90), Proteus sp (10), Pseudomonas sp (90), Serratia sp (20), and Staphylococcus sp (80). Higher resistance to beta-lactams was observed in the genera Pseudomonas, Escherichia and Klebsiella. For fluoroquinolones we found no significant resistance, with the exception of the genus Klebsiella. The most effective antibiotics were cephalosporins of the second and third generations, fluoroquinolones, and non-classical beta-lactams (cephamycins, moxalactam and monobactams). On the contrary, a pronounced resistance was found to penicillin, oxacillin, ticarcillin, ampicillin, methicillin, nalidixic acid and cinoxacin. These resistance patterns correspond to the high consumption of these antibiotics throughout the country.

  17. Discovery of a novel, potent and orally active series of gamma-lactams as selective NK1 antagonists.

    PubMed

    Paliwal, Sunil; Reichard, Gregory A; Shah, Sapna; Wrobleski, Michelle Laci; Wang, Cheng; Stengone, Carmine; Tsui, Hon-Chung; Xiao, Dong; Duffy, Ruth A; Lachowicz, Jean E; Nomeir, Amin A; Varty, Geoffrey B; Shih, Neng-Yang

    2008-07-15

    Strategic replacement of the nitrogen of the lead compound 1 in the original cyclic urea series with a carbon resulted in the discovery of a novel, potent and orally more efficacious gamma-lactam series of selective NK(1) antagonists. Optimization of the lactam series culminated in the identification of compounds with high binding affinity and excellent oral CNS activity.

  18. Genome Sequence of the Multiple-β-Lactam-Antibiotic-Resistant Bacterium Acidovorax sp. Strain MR-S7.

    PubMed

    Miura, Takamasa; Kusada, Hiroyuki; Kamagata, Yoichi; Hanada, Satoshi; Kimura, Nobutada

    2013-06-27

    Acidovorax sp. strain MR-S7 was isolated from activated sludge in a treatment system for wastewater containing β-lactam antibiotic pollutants. Strain MR-S7 demonstrates multidrug resistance for various types of β-lactam antibiotics at high levels of MIC. The draft genome sequence clarified that strain MR-S7 harbors unique β-lactamase genes.

  19. Complex Regulation Pathways of AmpC-Mediated β-Lactam Resistance in Enterobacter cloacae Complex.

    PubMed

    Guérin, François; Isnard, Christophe; Cattoir, Vincent; Giard, Jean Christophe

    2015-12-01

    Enterobacter cloacae complex (ECC), an opportunistic pathogen causing numerous infections in hospitalized patients worldwide, is able to resist β-lactams mainly by producing the AmpC β-lactamase enzyme. AmpC expression is highly inducible in the presence of some β-lactams, but the underlying genetic regulation, which is intricately linked to peptidoglycan recycling, is still poorly understood. In this study, we constructed different mutant strains that were affected in genes encoding enzymes suspected to be involved in this pathway. As expected, the inactivation of ampC, ampR (which encodes the regulator protein of ampC), and ampG (encoding a permease) abolished β-lactam resistance. Reverse transcription-quantitative PCR (qRT-PCR) experiments combined with phenotypic studies showed that cefotaxime (at high concentrations) and cefoxitin induced the expression of ampC in different ways: one involving NagZ (a N-acetyl-β-D-glucosaminidase) and another independent of NagZ. Unlike the model established for Pseudomonas aeruginosa, inactivation of DacB (also known as PBP4) was not responsible for a constitutive ampC overexpression in ECC, whereas it caused AmpC-mediated high-level β-lactam resistance, suggesting a post-transcriptional regulation mechanism. Global transcriptomic analysis by transcriptome sequencing (RNA-seq) of a dacB deletion mutant confirmed these results. Lastly, analysis of 37 ECC clinical isolates showed that amino acid changes in the AmpD sequence were likely the most crucial event involved in the development of high-level β-lactam resistance in vivo as opposed to P. aeruginosa where dacB mutations have been commonly found. These findings bring new elements for a better understanding of β-lactam resistance in ECC, which is essential for the identification of novel potential drug targets. PMID:26438498

  20. beta-Lactam resistance and beta-lactamases in bacteria of animal origin.

    PubMed

    Li, Xian-Zhi; Mehrotra, Manisha; Ghimire, Shiva; Adewoye, Lateef

    2007-04-15

    beta-Lactams are among the most clinically important antimicrobials in both human and veterinary medicine. Bacterial resistance to beta-lactams has been increasingly observed in bacteria, including those of animal origin. The mechanisms of beta-lactam resistance include inaccessibility of the drugs to their target, target alterations and/or inactivation of the drugs by beta-lactamases. The latter contributes predominantly to beta-lactam resistance in Gram-negative bacteria. A variety of beta-lactamases have been identified in bacteria derived from food-producing and companion animals and may further serve as a reservoir for beta-lactamase-producing bacteria in humans. While this review mainly describes beta-lactamases from animal-derived Escherichia coli and Salmonella spp., beta-lactamases from animal-derived Campylobacter spp., Enterococcus spp., Staphylococcus spp. and other pathogens are also discussed. Of particular concern are the increasingly-isolated plasmid-encoded AmpC-type CMY and extended-spectrum CTX-M beta-lactamases, which mediate acquired resistance to extended-spectrum beta-lactams. The genes encoding these enzymes often coexist with other antimicrobial resistance determinants and can also be associated with transposons/integrons, increasing the potential enrichment of multidrug resistant bacteria by multiple antimicrobial agents as well as dissemination of the resistance determinants among bacterial species. Characterization of beta-lactam-resistant animal-derived bacteria warrants further investigation of the type and distribution of beta-lactamases in bacteria of animal origin and their potential impact on human medicine.

  1. Complex Regulation Pathways of AmpC-Mediated β-Lactam Resistance in Enterobacter cloacae Complex

    PubMed Central

    Guérin, François; Isnard, Christophe; Giard, Jean Christophe

    2015-01-01

    Enterobacter cloacae complex (ECC), an opportunistic pathogen causing numerous infections in hospitalized patients worldwide, is able to resist β-lactams mainly by producing the AmpC β-lactamase enzyme. AmpC expression is highly inducible in the presence of some β-lactams, but the underlying genetic regulation, which is intricately linked to peptidoglycan recycling, is still poorly understood. In this study, we constructed different mutant strains that were affected in genes encoding enzymes suspected to be involved in this pathway. As expected, the inactivation of ampC, ampR (which encodes the regulator protein of ampC), and ampG (encoding a permease) abolished β-lactam resistance. Reverse transcription-quantitative PCR (qRT-PCR) experiments combined with phenotypic studies showed that cefotaxime (at high concentrations) and cefoxitin induced the expression of ampC in different ways: one involving NagZ (a N-acetyl-β-d-glucosaminidase) and another independent of NagZ. Unlike the model established for Pseudomonas aeruginosa, inactivation of DacB (also known as PBP4) was not responsible for a constitutive ampC overexpression in ECC, whereas it caused AmpC-mediated high-level β-lactam resistance, suggesting a post-transcriptional regulation mechanism. Global transcriptomic analysis by transcriptome sequencing (RNA-seq) of a dacB deletion mutant confirmed these results. Lastly, analysis of 37 ECC clinical isolates showed that amino acid changes in the AmpD sequence were likely the most crucial event involved in the development of high-level β-lactam resistance in vivo as opposed to P. aeruginosa where dacB mutations have been commonly found. These findings bring new elements for a better understanding of β-lactam resistance in ECC, which is essential for the identification of novel potential drug targets. PMID:26438498

  2. Mechanistic Insight into the Facilitation of β-Lactam Fragmentation through Metal Assistance.

    PubMed

    Casarrubios, Luis; Esteruelas, Miguel A; Larramona, Carmen; Lledós, Agustí; Muntaner, Jaime G; Oñate, Enrique; Ortuño, Manuel A; Sierra, Miguel A

    2015-11-16

    The mechanism of OsH6(PiPr3)2-mediated fragmentation of a 4-(2 pyridyl)-2-azetidinone has been investigated by DFT calculations. The addition of the C4-H bond of the substrate to OsH2(PiPr3)2 allows the active participation of an osmium lone pair in the B-type β-lactam fragmentation process. This new mechanism makes the N1-C4/C2-C3 fragmentation of the lactamic core thermally accessible through a stepwise process.

  3. Diastereoselective and enantioselective conjugate addition reactions utilizing α,β-unsaturated amides and lactams.

    PubMed

    Byrd, Katherine M

    2015-01-01

    The conjugate addition reaction has been a useful tool in the formation of carbon-carbon bonds. The utility of this reaction has been demonstrated in the synthesis of many natural products, materials, and pharmacological agents. In the last three decades, there has been a significant increase in the development of asymmetric variants of this reaction. Unfortunately, conjugate addition reactions using α,β-unsaturated amides and lactams remain underdeveloped due to their inherently low reactivity. This review highlights the work that has been done on both diastereoselective and enantioselective conjugate addition reactions utilizing α,β-unsaturated amides and lactams.

  4. Determination of chromatographic and spectrophotometric dissociation constants of some beta lactam antibiotics.

    PubMed

    Demiralay, Ebru Çubuk; Koç, Duygu; Daldal, Y Doğan; Cakır, Cansel

    2012-12-01

    In this work, dissociation constants values of seven beta lactam antibiotics in water and acetonitrile-water mixtures using spectrophotometric and reversed phase liquid chromatography methods were determined. The dissociation constant values of these compounds were calculated by NLREG and STAR programs. Aqueous pK(a) values of beta lactam antibiotics were calculated with extrapolation by means of the Yasuda-Shedlovsky and mole fraction equations. Finally, application of the different techniques was compared to the determination of aqueous pK(a) values of investigated compounds.

  5. Mechanistic Insight into the Facilitation of β-Lactam Fragmentation through Metal Assistance.

    PubMed

    Casarrubios, Luis; Esteruelas, Miguel A; Larramona, Carmen; Lledós, Agustí; Muntaner, Jaime G; Oñate, Enrique; Ortuño, Manuel A; Sierra, Miguel A

    2015-11-16

    The mechanism of OsH6(PiPr3)2-mediated fragmentation of a 4-(2 pyridyl)-2-azetidinone has been investigated by DFT calculations. The addition of the C4-H bond of the substrate to OsH2(PiPr3)2 allows the active participation of an osmium lone pair in the B-type β-lactam fragmentation process. This new mechanism makes the N1-C4/C2-C3 fragmentation of the lactamic core thermally accessible through a stepwise process. PMID:26437692

  6. Recent applications of ring-closing metathesis in the synthesis of lactams and macrolactams.

    PubMed

    Hassan, Hani Mutlak A

    2010-12-28

    Lactams are an important class of compounds owing to their presence in numerous biologically active molecules of natural and unnatural nature. They are also highly versatile intermediates that can be elaborated into interesting compounds for potential use in organic and medicinal chemistry endeavors. In this feature article, the reader will be given a background to olefin metathesis followed by concise discussions (with selected examples) to report recent applications of ring-closing metathesis to form lactams and macrolactams from acyclic diene precursors, an area which continues to deposit attractive applications in the chemical literature en route or in the final step to the target molecules.

  7. Diastereoselective and enantioselective conjugate addition reactions utilizing α,β-unsaturated amides and lactams

    PubMed Central

    2015-01-01

    Summary The conjugate addition reaction has been a useful tool in the formation of carbon–carbon bonds. The utility of this reaction has been demonstrated in the synthesis of many natural products, materials, and pharmacological agents. In the last three decades, there has been a significant increase in the development of asymmetric variants of this reaction. Unfortunately, conjugate addition reactions using α,β-unsaturated amides and lactams remain underdeveloped due to their inherently low reactivity. This review highlights the work that has been done on both diastereoselective and enantioselective conjugate addition reactions utilizing α,β-unsaturated amides and lactams. PMID:25977728

  8. Ab initio study of β-lactam antibiotics. I. Potential energy surface for the amidic CN bond breaking in the β-lactam + OH - reaction

    NASA Astrophysics Data System (ADS)

    Petrongolo, Carlo; Ranghino, Graziella; Scordamaglia, Raimondo

    1980-01-01

    The potential energy surface of the β-lactam + OH - reaction, related to the mode of action of β-lactam antibiotics, was investigated using the ab initio Hartree—Fock method with the STO-3G basis set. Three possible reaction paths for the B A C2 breaking of the amidic CN bond were obtained and discussed. The minimum-energy reaction path is characterized by the following processes: (1) the formation of a tetrahedral intermediate, ≈ 121 kcal mol -1 more stable than the reagents; (2) a barrier, ≈ 15 kcal mol -1 above the intermediate, which is mainly due to the partial breaking of the amidic bond; (3) the complete breaking of the amidic bond concerted with a proton transfer till the formation of the final product, ≈ 34 kcal mol -1 more stable than the intermediate. The evolution of some molecular orbitals and of the electron population along the reaction path was also discussed.

  9. Soluble penicillin-binding protein 2a: beta-lactam binding and inhibition by non-beta-lactams using a 96-well format.

    PubMed

    Toney, J H; Hammond, G G; Leiting, B; Pryor, K D; Wu, J K; Cuca, G C; Pompliano, D L

    1998-01-01

    High level methicillin resistance in Staphylococcus aureus is dependent upon the acquisition of the mecA gene encoding penicillin-binding protein 2a (PBP2a). PBP2a is a member of a family of peptidoglycan biosynthetic enzymes involved in assembly of the cell wall in bacteria and is poorly inactivated by beta-lactam antibiotics. We describe a 96-well-filter binding assay using recombinant, soluble PBP2a which allows for kinetic measurement of penicillin binding. The deacylation rate constant for the PBP2a-penicillin G covalent complex was found to be 5.7 +/- 1.0 x 10(-5) s-1 at 30 degrees C (half-life of approximately 200 min). For the PBP2a acylation reaction, the value of K(m) (penicillin G) = 0.5 +/- 0.1 mM and kcat = 1 x 10(-3) s-1, which yields a second-order rate constant (kcat/K(m)) for inactivation of 2.0 M-1 s-1. Using this assay, several non-beta-lactam inhibitors including Cibacron blue have been found which exhibit IC50 values between 10 and 30 microM. The binding affinities of several carbapenems and beta-lactams correlated well between the filter binding assay described in this report and an electrophoretic assay for PBP2a using membranes prepared form methicillin-resistant S. aureus. PMID:9448849

  10. Novel amino-β-lactam derivatives as potent cholesterol absorption inhibitors

    PubMed Central

    Dražić, Tonko; Molčanov, Krešimir; Sachdev, Vinay; Malnar, Martina; Hećimović, Silva; Patankar, Jay V.; Obrowsky, Sascha; Levak-Frank, Sanja; Habuš, Ivan; Kratky, Dagmar

    2014-01-01

    Two new trans-(3R,4R)-amino-β-lactam derivatives and their diastereoisomeric mixtures were synthesized as ezetimibe bioisosteres and tested in in vitro and in vivo experiments as novel β-lactam cholesterol absorption inhibitors. Both compounds exhibited low cytotoxicity in MDCKII, hNPC1L1/MDCKII, and HepG2 cell lines and potent inhibitory effect in hNPC1L1/MDCKII cells. In addition, these compounds markedly reduced cholesterol absorption in mice, resulting in reduced cholesterol concentrations in plasma, liver, and intestine. We determined the crystal structure of one amino-β-lactam derivative to establish unambiguously both the absolute and relative configuration at the new stereogenic centre C17, which was assigned to be S. The pKa values for both compounds are 9.35, implying that the amino-β-lactam derivatives and their diastereoisomeric mixtures are in form of ammonium salt in blood and the intestine. The IC50 value for the diastereoisomeric mixture is 60 μM. In vivo, it efficiently inhibited cholesterol absorption comparable to ezetimibe. PMID:25305716

  11. Copper(II) Carboxylate Promoted Intramolecular Carboamination of Alkenes for the Synthesis of Polycyclic Lactams

    PubMed Central

    Fuller, Peter H.; Chemler, Sherry R.

    2008-01-01

    The copper(II) carboxylate promoted intramolecular carboamination reactions of variously substituted γ-alkenyl amides have been investigated. These oxidative cyclization reactions efficiently provide polycyclic lactams, useful intermediates in nitrogen heterocycle synthesis, in good to excellent yields. The efficiency of the carboamination process is dependent upon the structure of the amide backbone as well as the nitrogen substituent. PMID:18044907

  12. Antimicrobial susceptibility and mechanisms of resistance to quinolones and beta-lactams in Acinetobacter genospecies 3.

    PubMed

    Ribera, A; Fernández-Cuenca, F; Beceiro, A; Bou, G; Martínez-Martínez, L; Pascual, A; Cisneros, J M; Rodríguez-Baño, J; Pachón, J; Vila, J

    2004-04-01

    Antimicrobial susceptibility was determined in 15 epidemiologically unrelated clinical isolates of Acinetobacter genospecies 3. Moreover, the mechanisms of resistance to some beta-lactam antibiotics may be associated with the presence of a chromosomal cephalosporinase, AmpC, and the resistance to quinolones related to mutations in the gyrA and parC genes.

  13. Antimicrobial Susceptibility and Mechanisms of Resistance to Quinolones and β-Lactams in Acinetobacter Genospecies 3

    PubMed Central

    Ribera, A.; Fernández-Cuenca, F.; Beceiro, A.; Bou, G.; Martínez-Martínez, L.; Pascual, A.; Cisneros, J. M.; Rodríguez-Baño, J.; Pachón, J.; Vila, J.

    2004-01-01

    Antimicrobial susceptibility was determined in 15 epidemiologically unrelated clinical isolates of Acinetobacter genospecies 3. Moreover, the mechanisms of resistance to some β-lactam antibiotics may be associated with the presence of a chromosomal cephalosporinase, AmpC, and the resistance to quinolones related to mutations in the gyrA and parC genes. PMID:15047561

  14. Amide and amine nucleophiles in polar radical crossover cycloadditions: synthesis of γ-lactams and pyrrolidines.

    PubMed

    Gesmundo, Nathan J; Grandjean, Jean-Marc M; Nicewicz, David A

    2015-03-01

    In this work we present a direct catalytic synthesis of γ-lactams and pyrrolidines from alkenes and activated unsaturated amides or protected unsaturated amines, respectively. Using a mesityl acridinium single electron photooxidant and a thiophenol cocatalyst under irradiation, we are able to directly forge these important classes of heterocycles with complete regiocontrol. PMID:25695366

  15. Simple and suitable immunosensor for β-lactam antibiotics analysis in real matrixes: milk, serum, urine.

    PubMed

    Merola, Giovanni; Martini, Elisabetta; Tomassetti, Mauro; Campanella, Luigi

    2015-03-15

    The anti-penicillin G was conjugated to avidin-peroxidase and biotin to obtain immunogen and competitor which were then used to develop a competitive immunosensor assay for the detection of penicillin G and other β-lactam antibiotics, with Kaff values of the order of 10(8) M(-1). The new immunosensor appears to afford a number of advantages in terms of sensitivity, possibility of "in situ" analysis, but especially of simplicity and lower costs, compared with other existing devices, or different chemical instrumental methods reported in the literature and used for the analysis of β-lactam compounds. Satisfactory results were found in the analysis of real matrixes and good recoveries were obtained by applying the standard addition method to spiked milk, urine, serum and drug samples. The new device uses an amperometric electrode for hydrogen peroxide as transducer, the BSA-penicillin G immobilized on polymeric membrane overlapping the amperometric transducer and the peroxidase enzyme as marker. It proved to be highly sensitive, inexpensive and easily reproducible; LOD was of the order of 10(-11)M. Lastly, the new immunosensor displayed low selectivity versus the entire class of β-lactam antibiotics and higher selectivity toward other classes of non-β-lactam antibiotics.

  16. Selective Pharmacologic Inhibition of a PASTA Kinase Increases Listeria monocytogenes Susceptibility to β-Lactam Antibiotics

    PubMed Central

    Pensinger, Daniel A.; Aliota, Matthew T.; Schaenzer, Adam J.; Boldon, Kyle M.; Ansari, Israr-ul H.; Vincent, William J. B.; Knight, Benjamin; Reniere, Michelle L.; Striker, Rob

    2014-01-01

    While β-lactam antibiotics are a critical part of the antimicrobial arsenal, they are frequently compromised by various resistance mechanisms, including changes in penicillin binding proteins of the bacterial cell wall. Genetic deletion of the penicillin binding protein and serine/threonine kinase-associated protein (PASTA) kinase in methicillin-resistant Staphylococcus aureus (MRSA) has been shown to restore β-lactam susceptibility. However, the mechanism remains unclear, and whether pharmacologic inhibition would have the same effect is unknown. In this study, we found that deletion or pharmacologic inhibition of the PASTA kinase in Listeria monocytogenes by the nonselective kinase inhibitor staurosporine results in enhanced susceptibility to both aminopenicillin and cephalosporin antibiotics. Resistance to vancomycin, another class of cell wall synthesis inhibitors, or antibiotics that inhibit protein synthesis was unaffected by staurosporine treatment. Phosphorylation assays with purified kinases revealed that staurosporine selectively inhibited the PASTA kinase of L. monocytogenes (PrkA). Importantly, staurosporine did not inhibit a L. monocytogenes kinase without a PASTA domain (Lmo0618) or the PASTA kinase from MRSA (Stk1). Finally, inhibition of PrkA with a more selective kinase inhibitor, AZD5438, similarly led to sensitization of L. monocytogenes to β-lactam antibiotics. Overall, these results suggest that pharmacologic targeting of PASTA kinases can increase the efficacy of β-lactam antibiotics. PMID:24867981

  17. Assays for therapeutic drug monitoring of β-lactam antibiotics: A structured review.

    PubMed

    Carlier, Mieke; Stove, Veronique; Wallis, Steven C; De Waele, Jan J; Verstraete, Alain G; Lipman, Jeffrey; Roberts, Jason A

    2015-10-01

    In some patient groups, including critically ill patients, the pharmacokinetics of β-lactam antibiotics may be profoundly disturbed due to pathophysiological changes in distribution and elimination. Therapeutic drug monitoring (TDM) is a strategy that may help to optimise dosing. The aim of this review was to identify and analyse the published literature on the methods used for β-lactam quantification in TDM programmes. Sixteen reports described methods for the simultaneous determination of three or more β-lactam antibiotics in plasma/serum. Measurement of these antibiotics, due to low frequency of usage relative to some other tests, is generally limited to in-house chromatographic methods coupled to ultraviolet or mass spectrometric detection. Although many published methods state they are fit for TDM, they are inconvenient because of intensive sample preparation and/or long run times. Ideally, methods used for routine TDM should have a short turnaround time (fast run-time and fast sample preparation), a low limit of quantification and a sufficiently high upper limit of quantification. The published assays included a median of 6 analytes [interquartile range (IQR) 4-10], with meropenem and piperacillin being the most frequently measured β-lactam antibiotics. The median run time was 8 min (IQR 5.9-21.3 min). There is also a growing number of methods measuring free concentrations. An assay that measures antibiotics without any sample preparation would be the next step towards real-time monitoring; no such method is currently available.

  18. Metal-containing Complexes of Lactams, Imidazoles, and Benzimidazoles and Their Biological Activity

    NASA Astrophysics Data System (ADS)

    Kukalenko, S. S.; Bovykin, B. A.; Shestakova, S. I.; Omel'chenko, A. M.

    1985-07-01

    The results of the latest investigations of the problem of the synthesis of metal-containing complexes of lactams, imidazoles, and benzimidazoles, their structure, and their stability in solutions are surveyed. Some data on their biological activity (pesticide and pharmacological) and the mechanism of their physiological action are presented. The bibliography includes 190 references.

  19. Δ-lactam derivative from thermophilic soil fungus exhibits in vitro anti-allergic activity.

    PubMed

    Andrioli, W J; Santos, M S; Silva, V B; Oliveira, R B; Chagas-Paula, D A; Jorge, J A; Furtado, N A J C; Pupo, M T; Silva, C H T P; Naal, R M Z G; Bastos, J K

    2012-01-01

    From cultures of thermophilic soil fungus Humicola grisea var thermoidea, a δ-lactam derivative (3-(2-(4-hydroxyphenyl)-2-oxoethyl)-5,6-dihydropyridin-2(1H)-one) that displayed anti-allergic activity was isolated, which was predicted by in silico computational chemistry approaches. The in vitro anti-allergic activity was investigated by β-hexosaminidase release assay in rat basophilic leukaemia RBL-2H3 cells. The δ-lactam derivative exhibited similar anti-allergic activity (IC(50) = 18.7 ± 6.7 µM) in comparison with ketotifen fumarate (IC(50) = 15.0 ± 1.3 µM) and stronger anti-allergic activity than azelastine (IC(50) = 32.0 µM). Also, the MTT cytotoxicity assay with RBL-2H3 cells showed that δ-lactam does not display cytotoxicity at concentrations lower than 50 µM. This study suggests that the δ-lactam derivative has the potential to be used as a lead compound in the development of anti-allergic drugs for clinical use in humans. PMID:22239222

  20. How β-Lactam Antibiotics Enter Bacteria: A Dialogue with the Porins

    PubMed Central

    Molitor, Alexander; Bolla, Jean-Michel; Bessonov, Andrey N.; Winterhalter, Mathias; Pagès, Jean-Marie

    2009-01-01

    Background Multi-drug resistant (MDR) infections have become a major concern in hospitals worldwide. This study investigates membrane translocation, which is the first step required for drug action on internal bacterial targets. β-lactams, a major antibiotic class, use porins to pass through the outer membrane barrier of Gram-negative bacteria. Clinical reports have linked the MDR phenotype to altered membrane permeability including porin modification and efflux pump expression. Methodology/Principal Findings Here influx of β-lactams through the major Enterobacter aerogenes porin Omp36 is characterized. Conductance measurements through a single Omp36 trimer reconstituted into a planar lipid bilayer allowed us to count the passage of single β-lactam molecules. Statistical analysis of each transport event yielded the kinetic parameters of antibiotic travel through Omp36 and distinguishable translocation properties of β-lactams were quantified for ertapenem and cefepime. Expression of Omp36 in an otherwise porin-null bacterial strain is shown to confer increases in the killing rate of these antibiotics and in the corresponding bacterial susceptibility. Conclusions/Significance We propose the idea of a molecular “passport” that allows rapid transport of substrates through porins. Deciphering antibiotic translocation provides new insights for the design of novel drugs that may be highly effective at passing through the porin constriction zone. Such data may hold the key for the next generation of antibiotics capable of rapid intracellular accumulation to circumvent the further development MDR infections. PMID:19434239

  1. Administration of parenteral prophylactic beta-lactam antibiotics in 2014: a review.

    PubMed

    Gordon, Ronald J

    2015-04-01

    The role of the anesthesiologist in reducing the incidence of surgical-site infections by the administration of prophylactic parenteral beta-lactam antibiotics is reviewed. Suggestions are made with regard to timing, dosing, and method of administration of these drugs to potentially reduce the risk of surgical-site infection.

  2. Epimerization and substrate gating by a TE domain in β-lactam antibiotic biosynthesis

    PubMed Central

    Gaudelli, Nicole M.; Townsend, Craig A.

    2014-01-01

    Nonribosomal peptide synthetases (NRPSs) are versatile engines of bioactive natural product biosynthesis that function according to the multiple carrier thiotemplate mechanism. C-terminal thioesterase (TE) domains of these giant modular proteins typically catalyze product release by hydrolysis or macrocylization. We now report an unprecedented, dual-function TE involved in nocardicin A biosynthesis, the paradigm monocyclic β-lactam antibiotic. Contrary to expectation, a stereodefined series of potential peptide substrates for the nocardicin TE domain failed to undergo hydrolysis. The stringent discrimination against peptide intermediates was dramatically overcome by prior monocyclic β-lactam formation at an L-seryl site. Kinetic data are interpreted such that the TE domain acts as a gatekeeper to hold the assembling peptide on an upstream domain until β-lactam formation takes place and then rapidly catalyzes epimerization, not previously observed as a TE catalytic function, and thioesterase cleavage to discharge a fully fledged pentapeptide β-lactam harboring nocardicin G, the universal precursor of the nocardicins. PMID:24531841

  3. β-Lactam formation by a non-ribosomal peptide synthetase during antibiotic biosynthesis

    PubMed Central

    Gaudelli, Nicole M.; Long, Darcie H.; Townsend, Craig A.

    2014-01-01

    Non-ribosomal peptide synthetases (NRPSs) are giant enzymes comprised of modules that house repeated sets of functional domains, which select, activate and couple amino acids drawn from a pool of nearly 500 potential building blocks.1 The structurally and stereochemically diverse peptides generated in this manner underlie the biosynthesis of a large sector of natural products. Many of their derived metabolites are bioactive such as the antibiotics vancomycin, bacitracin, daptomycin and the β-lactam-containing penicillins, cephalosporins and nocardicins. Although penicillins and cephalosporins are synthesised from a classically derived NRPS tripeptide (from ACVS, δ-(L-α-aminoadipyl)–L-cysteinyl–D-valine synthetase)2, we now report an unprecedented NRPS activity to both assemble a serine-containing peptide and mediate its cyclisation to the critical β-lactam ring of the nocardicin family of antibiotics. A histidine-rich condensation (C) domain, which typically carries out peptide bond formation during product assembly, was found to also synthesise the embedded 4-membered ring. Here, a mechanism is proposed and supporting experiments are described, which is distinct from the pathways that have evolved to the three other β-lactam antibiotic families: penicillin/cephalosporins, clavams and carbapenems. These findings raise the possibility that β-lactam rings can be regio- and stereospecifically integrated into engineered peptides for application as, for example, targeted protease inactivators.3,4 PMID:25624104

  4. Loss of a Class A Penicillin-Binding Protein Alters β-Lactam Susceptibilities in Mycobacterium tuberculosis.

    PubMed

    Wivagg, Carl N; Wellington, Samantha; Gomez, James E; Hung, Deborah T

    2016-02-12

    Recent studies have renewed interest in β-lactam antibiotics as a potential treatment for Mycobacterium tuberculosis infection. To explore the opportunities and limitations of this approach, we sought to better understand potential resistance mechanisms to β-lactam antibiotics in M. tuberculosis. We identified mutations in the penicillin-binding protein (PBP) ponA2 that were able to confer some degree of resistance to the cephalosporin subclass of β-lactams. Surprisingly, deletion of ponA2 also confers resistance, demonstrating that β-lactam resistance can spontaneously arise from PBP loss of function. We show that ponA2 mutants resistant to the cephalosporin subclass of β-lactams in fact show increased susceptibility to meropenem, a carbapenem that is known to target l,d-transpeptidases, thereby suggesting that in the absence of PonA2, an alternative mode of peptidoglycan synthesis likely becomes essential. Consistent with this hypothesis, a negative genetic selection identified the l,d-transpeptidase ldtMt2 as essential in the absence of ponA2. The mechanism of β-lactam resistance we outline is consistent with emerging models of β-lactam killing, while the investigation of ponA2 downstream and synthetic lethal genes sheds light on the mechanism of cell wall biosynthesis and the interaction between conventional PBPs and l,d-transpeptidases. PMID:27624961

  5. NADPH-dependent glutamate dehydrogenase in Penicillium chrysogenum is involved in regulation of beta-lactam production.

    PubMed

    Thykaer, Jette; Rueksomtawin, Kanchana; Noorman, Henk; Nielsen, Jens

    2008-04-01

    The interactions between the ammonium assimilatory pathways and beta-lactam production were investigated by disruption of the NADPH-dependent glutamate dehydrogenase gene (gdhA) in two industrial beta-lactam-producing strains of Penicillium chrysogenum. The strains used were an adipoyl-7-ADCA- and a penicillin-producing strain. The gdhA gene disruption caused a decrease in maximum specific growth rate of 26 % and 35 % for the adipoyl-7-ADCA-producing strain and the penicillin-producing strain, respectively, compared to the corresponding reference strains. Interestingly, no beta-lactam production was detected in either of the DeltagdhA strains. Supplementation with glutamate restored growth but no beta-lactam production was detected for the constructed strains. Cultures with high ammonium concentrations (repressing conditions) and with proline as nitrogen source (de-repressed conditions) showed continued beta-lactam production for the reference strains whereas the DeltagdhA strains remained non-productive under all conditions. By overexpressing the NAD-dependent glutamate dehydrogenase, the specific growth rate could be restored, but still no beta-lactam production was detected. The results indicate that the NADPH-dependent glutamate dehydrogenase may be directly or indirectly involved in the regulation of beta-lactam production in industrial strains of P. chrysogenum.

  6. Reduced Susceptibility to Extended-Spectrum β-Lactams in Vibrio cholerae Isolated in Bangladesh

    PubMed Central

    Ceccarelli, Daniela; Alam, Munirul; Huq, Anwar; Colwell, Rita R.

    2016-01-01

    β-lactams are antibiotic molecules able to inhibit cell wall biosynthesis. Among other mechanisms, resistance in Gram-negative bacteria is mostly associated with production of β-lactamase enzymes able to bind and hydrolyze the β-lactam ring. Extended-spectrum β-lactamases extend this ability also to third- and fourth-generation cephalosporins, as well as to carbapenems and monobactams. Vibrio cholerae is the causative agent of epidemic cholera and a public health burden for developing countries like Bangladesh. Although appropriate oral or intravenous rehydration is the therapy of choice for cholera, severe infections and V. cholerae-associated septicemia are treated with antimicrobial drugs, including doxycycline, erythromycin, azithromycin, ciprofloxacin, and/or third-generation cephalosporins. In the years after the introduction of antibiotics in clinical practice, V. cholerae developed resistance to commonly used drugs worldwide mostly through gene acquisition via horizontal gene transfer. Reduced susceptibility of V. cholerae to third-generation cephalosporins has been occasionally documented. However, carbapenemase-producing V. cholerae has been reported at higher rates than resistance to extended-spectrum β-lactams, mainly associated with blaNDM-1 emergence and successful plasmid dissemination. Recent findings suggest limited β-lactam resistance is present in V. cholerae O1 isolates collected during ecological and epidemiological surveillance in Bangladesh. However, a trend to intermediate-susceptibility insurgence was observed. Horizontal gene transfer of β-lactam resistance from enteric pathogens to environmental microorganisms should not be underrated, given the ability of V. cholerae to acquire new genetic information. PMID:27803895

  7. Physician approaches to beta-lactam use in patients with penicillin hypersensitivity.

    PubMed

    Prematta, Tracy; Shah, Shenil; Ishmael, Faoud T

    2012-01-01

    Beta-lactam antibiotics are widely used, but hypersensitivity reactions are common and difficult to manage. This study was designed to identify lack of knowledge regarding the safe use of alternative beta-lactams in penicillin-allergic patients and assess management differences between allergists and nonallergists. An electronic physician survey was sent to 623 providers in allergy, internal medicine, pediatrics, and family medicine, querying beta-lactam use in patients with a history of penicillin allergy. A total of 110 (17.7%) surveys were completed. For patients with a prior maculopapular rash to penicillin, most providers were uncomfortable prescribing penicillins again, although they would use other beta-lactams. In patients with an exfoliative dermatitis to penicillin, 46% of responders would not prescribe any beta-lactam again. For patients with a positive skin test to penicillin, only 45.1% of nonallergists were comfortable prescribing monobactams versus 62.5% of allergists; 30.3% of all responders would give a carbapenem. In patients with urticaria to penicillin, pediatricians were the most comfortable prescribing third- or fourth-generation cephalosporins. Providers (both allergists and nonallergists) were unfamiliar with the safety of prescribing penicillin in patients with history of maculopapular rash, the safety of monobactams, and low cross-reactivity with carbapenems in penicillin-allergic individuals. Nonallergists were also unfamiliar with the usefulness of penicillin skin testing. Improved education is needed to address these areas. Additionally, we found variability in responses regarding exfoliative dermatitis and comfort prescribing cephalosporins in patients with suspected IgE-mediated drug allergy to penicillin, highlighting the need for additional research in these areas.

  8. T3P-Promoted, Mild, One-Pot Syntheses of Constrained Polycyclic Lactam Dipeptide Analogues via Stereoselective Pictet-Spengler and Meyers Lactamization Reactions.

    PubMed

    Jida, Mouhamad; Van der Poorten, Olivier; Guillemyn, Karel; Urbanczyk-Lipkowska, Zofia; Tourwé, Dirk; Ballet, Steven

    2015-09-18

    A new convenient, mild, one-pot procedure is described for the diastereoselective synthesis of constrained 7,5- and 7,6-fused azabicycloalkanes. Using 2-formyl-L-tryptophan and 2-formyl-l-phenylalanine as bielectrophilic building blocks, T3P-mediated Pictet-Spengler and Meyers lactamization reactions were developed to present chiral and polycyclic aminoindolo- and aminobenzazepinone compounds in excellent yields. The conformationally constrained compounds can serve as templates for peptidomimetic research or polyheterocyclic privileged scaffolds. PMID:26322913

  9. Synthesis of ferrocene tethered open and macrocyclic bis-beta-lactams and bis-beta-amino acid derivatives.

    PubMed

    Sierra, Miguel A; Rodríguez-Fernández, Mamen; Casarrubios, Luis; Gómez-Gallego, Mar; Allen, Charles P; Mancheño, María José

    2009-10-21

    New bioorganometallic ferrocene derivatives are synthesized through a Diversity Oriented Synthesis strategy. Easily available ferrocene bisimines have been transformed into open ferrocenyl bis-beta-lactams. These compounds have demonstrated to be versatile synthons used in further transformations into new ferrocene bis-beta-amino acids. Carefully selected substituents submitted to ring closing metathesis (RCM) and Cu-catalyzed oxidative alkyne coupling conditions have also allowed the conversion of open substrates into ferrocenic macrocyclic bis-beta-lactams.

  10. A Potential Substrate Binding Conformation of β-Lactams and Insight into the Broad Spectrum of NDM-1 Activity

    PubMed Central

    Yuan, Qinghui; He, Lin

    2012-01-01

    New Delhi metallo-β-lactamase 1 (NDM-1) is a key enzyme that the pathogen Klebsiella pneumonia uses to hydrolyze almost all β-lactam antibiotics. It is currently unclear why NDM-1 has a broad spectrum of activity. Docking of the representatives of the β-lactam families into the active site of NDM-1 is reported here. All the β-lactams naturally fit the NDM-1 pocket, implying that NDM-1 can accommodate the substrates without dramatic conformation changes. The docking reveals two major binding modes of the β-lactams, which we tentatively name the S (substrate) and I (inhibitor) conformers. In the S conformers of all the β-lactams, the amide oxygen and the carboxylic group conservatively interact with two zinc ions, while the substitutions on the fused rings show dramatic differences in their conformations and positions. Since the bridging hydroxide ion/water in the S conformer is at the position for the nucleophilic attack, the S conformation may simulate the true binding of a substrate to NDM-1. The I conformer either blocks or displaces the bridging hydroxide ion/water, such as in the case of aztreonam, and is thus inhibitory. The docking also suggests that substitutions on the β-lactam ring are required for β-lactams to bind in the S conformation, and therefore, small β-lactams such as clavulanic acid would be inhibitors of NDM-1. Finally, our docking shows that moxalactam uses its tyrosyl-carboxylic group to compete with the S conformer and would thus be a poor substrate of NDM-1. PMID:22825119

  11. Antimicrobial activity of beta-lactams against multiresistant micro-organisms from the family Enterobacteriaceae, and genus Pseudomonas.

    PubMed

    Niebla, A; González, I; Vallín, C

    1994-01-01

    The antimicrobial activity of twenty beta-lactams was determined against multiresistant micro-organisms from the Enterobacteriaceae family (450) and the genus Pseudomonas (90). The antimicrobial susceptibility was assessed by the disk diffusion method. The most effective antibiotics were cephalosporins of the second and third generation, and non-classical beta-lactams (imipenem and moxalactam). A pronounced resistance was found to carbenicillin, ampicillin, cephalotin and cefazolin. These resistance patterns corresponded to a high consumption of these antibiotics.

  12. P4 capped amides and lactams as HCV NS3 protease inhibitors with improved potency and DMPK profile

    SciTech Connect

    Nair, Latha G.; Sannigrahi, Mousumi; Bogen, Stephane; Pinto, Patrick; Chen, Kevin X.; Prongay, Andrew; Tong, Xiao; Cheng, K.-C.; Girijavallabhann, Viyyoor; Njoroge, F. George

    2010-09-03

    SAR studies on the extension of P3 unit of Boceprevir (1, SCH 503034) with amides and lactams and their synthesis is described. Extensive SAR studies resulted in the identification of 36 bearing 4,4-dimethyl lactam as the new P4 cap unit with improved potency (K*{sub i}, EC 90 = 70 nM) and pharmacokinetic properties (Rat AUC (PO) = 3.52 {micro}M h) compared to 1.

  13. Penicillin-Binding Protein Transpeptidase Signatures for Tracking and Predicting β-Lactam Resistance Levels in Streptococcus pneumoniae

    PubMed Central

    Metcalf, Benjamin J.; Chochua, Sopio; Li, Zhongya; Gertz, Robert E.; Walker, Hollis; Hawkins, Paulina A.; Tran, Theresa; Whitney, Cynthia G.; McGee, Lesley; Beall, Bernard W.

    2016-01-01

    ABSTRACT β-Lactam antibiotics are the drugs of choice to treat pneumococcal infections. The spread of β-lactam-resistant pneumococci is a major concern in choosing an effective therapy for patients. Systematically tracking β-lactam resistance could benefit disease surveillance. Here we developed a classification system in which a pneumococcal isolate is assigned to a “PBP type” based on sequence signatures in the transpeptidase domains (TPDs) of the three critical penicillin-binding proteins (PBPs), PBP1a, PBP2b, and PBP2x. We identified 307 unique PBP types from 2,528 invasive pneumococcal isolates, which had known MICs to six β-lactams based on broth microdilution. We found that increased β-lactam MICs strongly correlated with PBP types containing divergent TPD sequences. The PBP type explained 94 to 99% of variation in MICs both before and after accounting for genomic backgrounds defined by multilocus sequence typing, indicating that genomic backgrounds made little independent contribution to β-lactam MICs at the population level. We further developed and evaluated predictive models of MICs based on PBP type. Compared to microdilution MICs, MICs predicted by PBP type showed essential agreement (MICs agree within 1 dilution) of >98%, category agreement (interpretive results agree) of >94%, a major discrepancy (sensitive isolate predicted as resistant) rate of <3%, and a very major discrepancy (resistant isolate predicted as sensitive) rate of <2% for all six β-lactams. Thus, the PBP transpeptidase signatures are robust indicators of MICs to different β-lactam antibiotics in clinical pneumococcal isolates and serve as an accurate alternative to phenotypic susceptibility testing. PMID:27302760

  14. Discovery of wall teichoic acid inhibitors as potential anti-MRSA β-lactam combination agents.

    PubMed

    Wang, Hao; Gill, Charles J; Lee, Sang H; Mann, Paul; Zuck, Paul; Meredith, Timothy C; Murgolo, Nicholas; She, Xinwei; Kales, Susan; Liang, Lianzhu; Liu, Jenny; Wu, Jin; Santa Maria, John; Su, Jing; Pan, Jianping; Hailey, Judy; Mcguinness, Debra; Tan, Christopher M; Flattery, Amy; Walker, Suzanne; Black, Todd; Roemer, Terry

    2013-02-21

    Innovative strategies are needed to combat drug resistance associated with methicillin-resistant Staphylococcus aureus (MRSA). Here, we investigate the potential of wall teichoic acid (WTA) biosynthesis inhibitors as combination agents to restore β-lactam efficacy against MRSA. Performing a whole-cell pathway-based screen, we identified a series of WTA inhibitors (WTAIs) targeting the WTA transporter protein, TarG. Whole-genome sequencing of WTAI-resistant isolates across two methicillin-resistant Staphylococci spp. revealed TarG as their common target, as well as a broad assortment of drug-resistant bypass mutants mapping to earlier steps of WTA biosynthesis. Extensive in vitro microbiological analysis and animal infection studies provide strong genetic and pharmacological evidence of the potential effectiveness of WTAIs as anti-MRSA β-lactam combination agents. This work also highlights the emerging role of whole-genome sequencing in antibiotic mode-of-action and resistance studies.

  15. How similar is the electronic structures of β-lactam and alanine?

    NASA Astrophysics Data System (ADS)

    Chatterjee, Subhojyoti; Ahmed, Marawan; Wang, Feng

    2016-02-01

    The C1s spectra of β-lactam i.e. 2-azetidinone (C3H5NO), a drug and L-alanine (C3H7NO2), an amino acid, exhibit striking similarities, which may be responsible for the competition between 2-azetidinone and the alanyl-alanine moiety in biochemistry. The present study is to reveal the degree of similarities and differences between their electronic structures of the two model molecular pairs. It is found that the similarities in C1s and inner valence binding energy spectra are due to their bonding connections but other properties such as ring structure (in 2-azetidinone) and chiral carbon (alanine) can be very different. Further, the inner valence region of ionization potential greater than 18 eV for 2-azetidinone and alanine is also significantly similar. Finally the strained lactam ring exhibits more chemical reactivity measured at all non-hydrogen atoms by Fukui functions with respect to alanine.

  16. The beta-lactam antibiotic, ceftriaxone, inhibits the development of opioid-induced hyperalgesia in mice.

    PubMed

    Chen, Zhijun; He, Ying; Wang, Zaijie Jim

    2012-02-16

    The glutamate transporter GLT-1 is primarily responsible for glutamate clearance in the spinal cord. beta-Lactam antibiotics have been shown to attenuate neuropathic pain behaviors by promoting GLT-1 expression and function in the CNS. The present study tested the hypothesis that ceftriaxone, a prototype beta-lactam antibiotic, can prevent the development of opioid-induced hyperalgesia (OIH) in mice. Repeated morphine administration produced mechanical allodynia and thermal hyperalgesia, signs of OIH, and reduced spinal GLT-1 expression in mice. Ceftriaxone (200mg/kg/d, i.p., for 7 d) inhibited OIH. Correlating with the behavioral effects, ceftriaxone reversed downregulation of GLT-1 expression that was induced by OIH. These results suggest that ceftriaxone inhibited the development of OIH by up-regulating spinal GLT-1 expression.

  17. Avibactam is a covalent, reversible, non–β-lactam β-lactamase inhibitor

    PubMed Central

    Ehmann, David E.; Jahić, Haris; Ross, Philip L.; Gu, Rong-Fang; Hu, Jun; Kern, Gunther; Walkup, Grant K.; Fisher, Stewart L.

    2012-01-01

    Avibactam is a β-lactamase inhibitor that is in clinical development, combined with β-lactam partners, for the treatment of bacterial infections comprising Gram-negative organisms. Avibactam is a structural class of inhibitor that does not contain a β-lactam core but maintains the capacity to covalently acylate its β-lactamase targets. Using the TEM-1 enzyme, we characterized avibactam inhibition by measuring the on-rate for acylation and the off-rate for deacylation. The deacylation off-rate was 0.045 min−1, which allowed investigation of the deacylation route from TEM-1. Using NMR and MS, we showed that deacylation proceeds through regeneration of intact avibactam and not hydrolysis. Other than TEM-1, four additional clinically relevant β-lactamases were shown to release intact avibactam after being acylated. We showed that avibactam is a covalent, slowly reversible inhibitor, which is a unique mechanism of inhibition among β-lactamase inhibitors. PMID:22753474

  18. Analysis of different beta-lactams antibiotics in pharmaceutical preparations using micellar electrokinetic capillary chromatography.

    PubMed

    Pérez, M I Bailón; Rodríguez, L Cuadros; Cruces-Blanco, C

    2007-01-17

    The potential of micellar electrokinetic capillary chromatography (MEKC) for analyzing nine beta-lactams antibiotics (cloxacillin, dicloxacillin, oxacillin, penicillin G, penicillin V, ampicillin, nafcillin, piperacillin, amoxicillin) in different pharmaceutical preparations, have been demonstrated. An experimental design strategy has been applied to optimize the main variables: pH and buffer concentration, concentration of the micellar medium, separation voltage and capillary temperature. Borate buffer (26mM) at pH 8.5 containing 100mM sodium dodecyl sulphate (SDS) was used as the background electrolyte. The method was validated. Linearity, limit of detection and quantitation and precision were established for each compound. The analysis of some of the beta-lactams in Orbenin capsules, Britapen tables and in Veterin-Micipen injectable, all used in human and veterinary medicine, have demonstrated the applicability of these technique for quality control in the pharmaceutical industry.

  19. Beta-lactam antibiotics induce a lethal malfunctioning of the bacterial cell wall synthesis machinery

    PubMed Central

    Cho, Hongbaek; Uehara, Tsuyoshi; Bernhardt, Thomas G.

    2014-01-01

    SUMMARY Penicillin and related beta-lactams comprise one of our oldest and most widely used antibiotic therapies. These drugs have long been known to target enzymes called penicillin-binding proteins (PBPs) that build the bacterial cell wall. Investigating the downstream consequences of target inhibition and how they contribute to the lethal action of these important drugs, we demonstrate that beta-lactams do more than just inhibit the PBPs as is commonly believed. Rather, they induce a toxic malfunctioning of their target biosynthetic machinery involving a futile cycle of cell wall synthesis and degradation, thereby depleting cellular resources and bolstering their killing activity. Characterization of this mode of action additionally revealed a quality-control function for enzymes that cleave bonds in the cell wall matrix. The results thus provide insight into the mechanism of cell wall assembly and suggest how best to interfere with the process for future antibiotic development. PMID:25480295

  20. Towards a general synthesis of 3-metal-substituted β-lactams.

    PubMed

    Baeza, Beatriz; Casarrubios, Luis; Sierra, Miguel A

    2013-08-26

    Joining metals and antibiotics: Studies towards a general method for the synthesis of β-lactams that have a metal complex moiety attached to the C3-position are reported (see scheme). The cis/trans selectivity of the reactions ranges from low in complexes containing the alkyne moiety joined directly to the cyclopentadienyl ring to complete when the metal moiety is separated from the reactive alkyne by an alkynyl-aryl fragment.

  1. beta-Lactam resistance phenotype determination in Escherichia coli isolates from University Malaya Medical Centre.

    PubMed

    Wong, Jeanne Sze Lyn; Mohd Azri, Zainal Abidin; Subramaniam, Geetha; Ho, Siaw Eng; Palasubramaniam, Selvi; Navaratnam, Parasakthi

    2003-12-01

    beta-Lactamases have been identified as the major cause of antimicrobial resistance to beta-lactam antibiotics in Escherichia coli. The activities of ampicillin-sulbactam and amoxicillin-clavulanate as well as a range of beta-lactam antibiotics were studied with 87 clinical E. coli isolates from patients of the University Malaya Medical Center using the disc diffusion technique. Susceptible, intermediate and resistant categories were established based on the diameter of zones of inhibition set by the National Committee for Clinical Laboratory Standards (NCCLS). The isolates were then classified into 6 phenotypes according to the criteria stated in the methodology: S (susceptible to all beta-lactams); TL (resistant to aminopenicillins; amoxicillin-clavulanate susceptible and susceptible or intermediate to ampicillin-sulbactam); TI (resistant to aminopenicillins and ampicillin-sulbactam; susceptible to amoxicilin-clavulanate); TH-IRT (resistant to aminopenicillins; intermediate or resistant to amoxicillin-clavulanate; resistant to ampicillin-sulbactam); ESBL (resistant to aminopenicillins and oxyimino cephalosporins; positive results with the double-disc diffusion test); and CP (resistant to aminopenicillins, beta-lactam-beta-lactamase inhibitor combinations, oxyimino cephalosporins and cephamycins). Results showed that the TL phenotype was the commonest (40.2% of the isolates) followed by S (31%), TH-IRT (16.1%), ESBL and CP (3.4% each) and TI (2.3%). One isolate showed both ESBL and CP phenotypes while two isolates were classified as inconclusive. Representatives from each phenotype were further analysed for the presence of beta-lactamases which revealed a predominance of TEM and SHV enzyme producers. PCR-SSCP analysis of the SHV gene from all the ESBL and CP isolates revealed the predominance of SHV 5-type enzyme which was concurrent with our previous studies.

  2. [Importance of quality control for the detection of β-lactam antibiotic resistance in Enterobacteriaceae].

    PubMed

    Rivera, Alba; Larrosa, Nieves; Mirelis, Beatriz; Navarro, Ferran

    2014-02-01

    β-lactam antimicrobial agents are frequently used to treat infections caused by Enterobacteriaceae. The main mechanism of resistance to these antibiotics is the production of certain enzymes, collectively named β-lactamases. Due to their substrate profile and their epidemiological implications, the most clinically important β-lactamases are extended-spectrum β-lactamases, class C β-lactamases and carbapenemases. Phenotypic detection of these enzymes may be complicated and is based on the use of specific inhibitors of each β-lactamase and on the loss of activity on some β-lactam indicators. Various international committees postulate that it is no longer necessary to interpret the susceptibility results or determine the mechanism of resistance. Several critics disagree, however, and consider that susceptibility results should be interpreted until more data are available on the clinical efficacy of treatment with β-lactams. Given these methodological difficulties and constant changes in the interpretation criteria, we consider that training and external quality controls are essential to keep updated in this field. For learning purposes, these external quality controls should always be accompanied by a review of the results and methodology used, and the analysis of errors. In this paper we review and contextualize all the aspects related to the detection and interpretation of these β-lactamases.

  3. Kinase Inhibitors that Increase the Sensitivity of Methicillin Resistant Staphylococcus aureus to β-Lactam Antibiotics

    PubMed Central

    Vornhagen, Jay; Burnside, Kellie; Whidbey, Christopher; Berry, Jessica; Qin, Xuan; Rajagopal, Lakshmi

    2015-01-01

    Staphylococcus aureus are Gram-positive bacteria that are the leading cause of recurrent infections in humans that include pneumonia, bacteremia, osteomyelitis, arthritis, endocarditis, and toxic shock syndrome. The emergence of methicillin resistant S. aureus strains (MRSA) has imposed a significant concern in sustained measures of treatment against these infections. Recently, MRSA strains deficient in expression of a serine/threonine kinase (Stk1 or PknB) were described to exhibit increased sensitivity to β-lactam antibiotics. In this study, we screened a library consisting of 280 drug-like, low-molecular-weight compounds with the ability to inhibit protein kinases for those that increased the sensitivity of wild-type MRSA to β-lactams and then evaluated their toxicity in mice. We report the identification of four kinase inhibitors, the sulfonamides ST085384, ST085404, ST085405, and ST085399 that increased sensitivity of WT MRSA to sub-lethal concentrations of β-lactams. Furthermore, these inhibitors lacked alerting structures commonly associated with toxic effects, and toxicity was not observed with ST085384 or ST085405 in vivo in a murine model. These results suggest that kinase inhibitors may be useful in therapeutic strategies against MRSA infections. PMID:26506394

  4. In vitro susceptibility of Yersinia kristensenii strains to beta-lactam antibiotics.

    PubMed

    Bejar, V; Calvo, C; Ramos Cormenzana, A

    1986-01-01

    The beta-lactam antibiotic susceptibility and beta-lactamase activity of 22 Yersinia kristensenii strains were determined in order to compare these properties with those reported for Y. enterocolitica. Carbenicillin, cephaloridine, cefoxitin and cefotaxime were the most active antimicrobial agents tested against Y. kristensenii. All strains were resistant to penicillin G, cephalothin, cephapirin and cephalexin. On the other hand, ampicillin and cefazolin activity could be considerated as intermediate. The influence of incubation temperature was also evaluated. All Y. kristensenii strains were more resistant at 22 degrees C than at 37 degrees C to beta-lactam antibiotics. Susceptibility to cefazolin was not affected by the incubation temperature. All Y. kristensenii strains produce beta-lactamase. The substrate profile of this enzyme is that of a cephalosporinase: its activity was highest on cephalothin, cephapirin, cephaloridine and cefazolin, while hydrolysis of ampicillin and carbenicillin was not detected. beta-Lactamase-resistant cefoxitin and cefotaxime were not hydrolysed. A correlation between beta-lactam antibiotic susceptibility and the hydrolysis rate of these antibiotics by beta-lactamase was observed, except for cephaloridine and cefazolin; these antibiotics were very good beta-lactamase substrates, but their MIC values were low.

  5. Bridgehead lithiation-substitution of bridged ketones, lactones, lactams, and imides: experimental observations and computational insights.

    PubMed

    Hayes, Christopher J; Simpkins, Nigel S; Kirk, Douglas T; Mitchell, Lee; Baudoux, Jerome; Blake, Alexander J; Wilson, Claire

    2009-06-17

    The viability of bridgehead lithiation-substitution of bridged carbonyl compounds has been tested in the laboratory, and the results were rationalized with the aid of a computational study. Lithiation-substitution was found to be possible for ketones, lactones, lactams, and imides having small bridges, including examples having [3.2.1], [3.2.2], [3.3.1], [4.2.1], and [4.3.1] skeletons. Smaller systems, where the sum of the bridging atoms (S) was 5, for example [2.2.1] or [3.1.1] ketones or [2.2.1] lactams, did not undergo controlled bridgehead substitution. Ketones or lactams having a [2.2.2] structure also did not give bridgehead substitution. B3LYP calculations accurately predict this behavior with negative DeltaE(rxn) values being calculated for the successful deprotonations and positive DeltaE(rxn) values being calculated for the unsuccessful ones. NBO calculations were also performed on the anionic deprotonated species, and these show that some structures are best represented as bridgehead enolates and some are best represented as alpha-keto carbanions. PMID:19507906

  6. Evaluation of nisin-β-lactam antibiotics against clinical strains of Salmonella enterica serovar Typhi.

    PubMed

    Rishi, Praveen; Preet Singh, Aman; Garg, Neha; Rishi, Madhuri

    2014-12-01

    There is an imperative need to discover novel antimicrobials and anti-infective agents and build up innovative strategies to combat multidrug-resistant Salmonella. In this context, we had earlier confirmed that nisin has the potential to act in conjunction with β-lactams against murine salmonellosis using standard strain. However, evaluation of efficacy of these combinations against clinical isolates of Salmonella could be the next key step to confirm the value added potential of this peptide. The present study was therefore planned to validate the synergistic effects of nisin-β-lactams combinations against clinical isolates of Salmonella enterica serovar Typhi. MICs of the selected β-lactams, EDTA and nisin were determined by micro and macro broth dilution assays. In-vitro synergism between the agents was evaluated by fractional inhibitory concentration (FIC) index (checkerboard test) and time-kill assay. All the tested combinations showed synergy against the tested clinical strains except three, as evident by FIC index (checkerboard test) and time-kill assay. Especially, nisin-ceftriaxone and nisin-cefotaxime combinations demonstrated excellent synergistic activity. In view of the encouraging results obtained from the previous and present study, further studies need to be carried out using large number of strains from different regions to take into account the clinical variability of the strains. Though it is a simple study but highlights an important point about the possibility of using the said combination while making strategies to develop newer formulations.

  7. Atomistic Model for the Polyamide Formation from β-Lactam Catalyzed by Candida Antarctica Lipase B

    SciTech Connect

    Baum, Iris; Elsasser, Brigitta M.; Schwab, Leendert; Loos, Katja; Fels, Gregor

    2011-04-01

    Candida antarctica lipase B (CALB) is an established biocatalyst for a variety of transesterification, amidation, and polymerization reactions. In contrast to polyesters, polyamides are not yet generally accessible via enzymatic polymerization. In this regard, an enzyme-catalyzed ring-opening polymerization of {beta}-lactam (2-azetidinone) using CALB is the first example of an enzymatic polyamide formation yielding unbranched poly({beta}-alanine), nylon 3. The performance of this polymerization, however, is poor, considering the maximum chain length of 18 monomer units with an average length of 8, and the molecular basis of the reaction so far is not understood. We have employed molecular modeling techniques using docking tools, molecular dynamics, and QM/MM procedures to gain insight into the mechanistic details of the various reaction steps involved. As a result, we propose a catalytic cycle for the oligomerization of {beta}-lactam that rationalizes the activation of the monomer, the chain elongation by additional {beta}-lactam molecules, and the termination of the polymer chain. In addition, the processes leading to a premature chain termination are studied. Particularly, the QM/MM calculation enables an atomistic description of all eight steps involved in the catalytic cycle, which features an in situ-generated {beta}-alanine as the elongating monomer and which is compatible with the experimental findings.

  8. Kinase Inhibitors that Increase the Sensitivity of Methicillin Resistant Staphylococcus aureus to β-Lactam Antibiotics.

    PubMed

    Vornhagen, Jay; Burnside, Kellie; Whidbey, Christopher; Berry, Jessica; Qin, Xuan; Rajagopal, Lakshmi

    2015-01-01

    Staphylococcus aureus are Gram-positive bacteria that are the leading cause of recurrent infections in humans that include pneumonia, bacteremia, osteomyelitis, arthritis, endocarditis, and toxic shock syndrome. The emergence of methicillin resistant S. aureus strains (MRSA) has imposed a significant concern in sustained measures of treatment against these infections. Recently, MRSA strains deficient in expression of a serine/threonine kinase (Stk1 or PknB) were described to exhibit increased sensitivity to β-lactam antibiotics. In this study, we screened a library consisting of 280 drug-like, low-molecular-weight compounds with the ability to inhibit protein kinases for those that increased the sensitivity of wild-type MRSA to β-lactams and then evaluated their toxicity in mice. We report the identification of four kinase inhibitors, the sulfonamides ST085384, ST085404, ST085405, and ST085399 that increased sensitivity of WT MRSA to sub-lethal concentrations of β-lactams. Furthermore, these inhibitors lacked alerting structures commonly associated with toxic effects, and toxicity was not observed with ST085384 or ST085405 in vivo in a murine model. These results suggest that kinase inhibitors may be useful in therapeutic strategies against MRSA infections. PMID:26506394

  9. Evaluation of nisin-β-lactam antibiotics against clinical strains of Salmonella enterica serovar Typhi.

    PubMed

    Rishi, Praveen; Preet Singh, Aman; Garg, Neha; Rishi, Madhuri

    2014-12-01

    There is an imperative need to discover novel antimicrobials and anti-infective agents and build up innovative strategies to combat multidrug-resistant Salmonella. In this context, we had earlier confirmed that nisin has the potential to act in conjunction with β-lactams against murine salmonellosis using standard strain. However, evaluation of efficacy of these combinations against clinical isolates of Salmonella could be the next key step to confirm the value added potential of this peptide. The present study was therefore planned to validate the synergistic effects of nisin-β-lactams combinations against clinical isolates of Salmonella enterica serovar Typhi. MICs of the selected β-lactams, EDTA and nisin were determined by micro and macro broth dilution assays. In-vitro synergism between the agents was evaluated by fractional inhibitory concentration (FIC) index (checkerboard test) and time-kill assay. All the tested combinations showed synergy against the tested clinical strains except three, as evident by FIC index (checkerboard test) and time-kill assay. Especially, nisin-ceftriaxone and nisin-cefotaxime combinations demonstrated excellent synergistic activity. In view of the encouraging results obtained from the previous and present study, further studies need to be carried out using large number of strains from different regions to take into account the clinical variability of the strains. Though it is a simple study but highlights an important point about the possibility of using the said combination while making strategies to develop newer formulations. PMID:24961707

  10. Synergistic, collaterally sensitive β-lactam combinations suppress resistance in MRSA

    PubMed Central

    Gonzales, Patrick R.; Pesesky, Mitchell W.; Bouley, Renee; Ballard, Anna; Biddy, Brent A.; Suckow, Mark A.; Wolter, William R.; Schroeder, Valerie A.; Burnham, Carey-Ann D.; Mobashery, Shahriar; Chang, Mayland; Dantas, Gautam

    2015-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most prevalent multidrug-resistant pathogens worldwide, exhibiting increasing resistance to the latest antibiotic therapies. Here we show that the triple β-lactam combination meropenem/piperacillin/tazobactam (ME/PI/TZ) acts synergistically and is bactericidal against MRSA N315 and 72 clinical MRSA isolates in vitro, and clears MRSA N315 infection in a mouse model. ME/PI/TZ suppresses evolution of resistance in MRSA via reciprocal collateral sensitivity of its constituents. We demonstrate that these activities also extend to other carbapenem/penicillin/β-lactamase inhibitor combinations. ME/PI/TZ circumvents the tight regulation of the mec and bla operons in MRSA, the basis for inducible resistance to β-lactam antibiotics. Furthermore, ME/PI/TZ subverts the function of penicillin-binding protein 2a (PBP2a) action via allostery, which we propose as the mechanism for both synergy and collateral sensitivity. Showing similar in vivo activity to linezolid, ME/PI/TZ demonstrates that combinations of older β-lactam antibiotics could be effective against MRSA infections in humans. PMID:26368589

  11. Insertion of multiple alpha-amino gamma-lactam (Agl) residues into a peptide sequence by solid-phase synthesis on synphase lanterns.

    PubMed

    Ronga, Luisa; Jamieson, Andrew G; Beauregard, Kim; Quiniou, Christiane; Chemtob, Sylvain; Lubell, William D

    2010-01-01

    The insertion of lactams into peptide analogs can enhance potency and improve receptor selectivity. The synthesis of lactam-bridged peptide sequences has been accomplished by a solid-phase approach on SynPhase lanterns using cyclic (R)- and (S)-oxathiazinane ester (2) to annulate the amino lactam residue onto the peptide chain. Parallel synthesis of alpha-amino gamma-lactam analogs of the allosteric modulator of IL-1 receptor 101.10 (D-Arg-D-Tyr-D-Thr-D-Val-D-Glu-D-Leu-D-Ala: rytvela) was performed by split-mix chemistry on the lanterns. In particular, the double insertion of alpha-amino gamma-lactams in the same peptide sequence has been accomplished by this effective method for the solid-supported combinatorial synthesis of lactam-bridged peptides. Peptides bearing an Agl residue exhibited curve shapes indicative of turn conformations in their circular dichroism spectra. PMID:20225301

  12. Noncovalent Interaction Energies in Covalent Complexes: TEM-1 beta-Lactamase and beta-Lactams

    SciTech Connect

    Wang, Xiaojun; Minasov, George; Shoichet, Brian K.

    2010-03-08

    The class A {beta}-lactamase TEM-1 is a key bacterial resistance enzyme against {beta}-lactam antibiotics, but little is known about the energetic bases for complementarity between TEM-1 and its inhibitors. Most inhibitors form a covalent adduct with the catalytic Ser70, making the measurement of equilibriumconstants, and hence interaction energies, technically difficult. This study evaluates noncovalent interactions withincovalent complexes by examining the differential stability of TEM-1 and its inhibitor adducts. The thermal denaturation of TEM-1 follows a two-state, reversible model with a melting temperature (T{sub m}) of 51.6 C and a van't Hoff enthalpy of unfolding ({Delta}H{sub VH}) of 146.2 kcal/mol at pH 7.0. The stability of the enzyme changes on forming an inhibitor adduct. As expected, some inhibitors stabilize TEM-1; transition-state analogues increase the T{sub m} by up to 3.7 C(1.7 kcal/mol). Surprisingly, all {beta}-lactam covalent acyl-enzyme complexes tested destabilize TEM-1 significantly relative to the apoenzyme. For instance, the clinically used inhibitor clavulanic acid and the {beta}-lactamase-resistant {beta}-lactams moxalactam and imipenem destabilize TEM-1 by over 2.6 C (1.2 kcal/mol) in their covalent adducts. Based on the structure of the TEM-1/imipenem complex (Maveyraud et al., J Am Chem Soc 1998;120:9748-52), destabilization by moxalactam and imipenem is thought to be caused by a steric clash between the side-chain of Asn132 and the 6(7)-{alpha} group of these {beta}-lactams. To test this hypothesis, the mutant enzyme N132A was made. In contrast with wild-type, the covalent complexes between N132A and both imipenem and moxalactam stabilize the enzyme, consistent with the hypothesis. To investigate the structural bases of this dramatic change instability, the structure of N132A/imipenem was determined by X-ray crystallography. In the complex with N132A, imipenemadopts a very different conformation from that observed in the wild

  13. Beta-lactam dosing in critically ill patients with septic shock and continuous renal replacement therapy

    PubMed Central

    2014-01-01

    Although early and appropriate antibiotic therapy remains the most important intervention for successful treatment of septic shock, data guiding optimization of beta-lactam prescription in critically ill patients prescribed with continuous renal replacement therapy (CRRT) are still limited. Being small hydrophilic molecules, beta-lactams are likely to be cleared by CRRT to a significant extent. As a result, additional variability may be introduced to the per se variable antibiotic concentrations in critically ill patients. This article aims to describe the current clinical scenario for beta-lactam dosing in critically ill patients with septic shock and CRRT, to highlight the sources of variability among the different studies that reduce extrapolation to clinical practice, and to identify the opportunities for future research and improvement in this field. Three frequently prescribed beta-lactams (meropenem, piperacillin and ceftriaxone) were chosen for review. Our findings showed that present dosing recommendations are based on studies with drawbacks limiting their applicability in the clinical setting. In general, current antibiotic dosing regimens for CRRT follow a one-size-fits-all fashion despite emerging clinical data suggesting that drug clearance is partially dependent on CRRT modality and intensity. Moreover, some studies pool data from heterogeneous populations with CRRT that may exhibit different pharmacokinetics (for example, admission diagnoses different to septic shock, such as trauma), which also limit their extrapolation to critically ill patients with septic shock. Finally, there is still no consensus regarding the %T>MIC (percentage of dosing interval when concentration of the antibiotic is above the minimum inhibitory concentration of the pathogen) value that should be chosen as the pharmacodynamic target for antibiotic therapy in patients with septic shock and CRRT. For empirically optimized dosing, during the first day a loading dose is required

  14. Exposure of Staphylococcus aureus to Subinhibitory Concentrations of β-Lactam Antibiotics Induces Heterogeneous Vancomycin-Intermediate Staphylococcus aureus

    PubMed Central

    Roch, Mélanie; Clair, Perrine; Renzoni, Adriana; Reverdy, Marie-Elisabeth; Dauwalder, Olivier; Bes, Michèle; Martra, Annie; Freydière, Anne-Marie; Laurent, Frédéric; Reix, Philippe; Dumitrescu, Oana

    2014-01-01

    Glycopeptides are known to select for heterogeneous vancomycin-intermediate Staphylococcus aureus (h-VISA) from susceptible strains. In certain clinical situations, h-VISA strains have been isolated from patients without previous exposure to glycopeptides, such as cystic fibrosis patients, who frequently receive repeated treatments with beta-lactam antibiotics. Our objective was to determine whether prolonged exposure to beta-lactam antibiotics can induce h-VISA. We exposed 3 clinical vancomycin-susceptible methicillin-resistant Staphylococcus aureus (MRSA) strains to ceftazidime, ceftriaxone, imipenem, and vancomycin (as a control) at subinhibitory concentrations for 18 days in vitro. Population analyses showed progressive increases in vancomycin resistance; seven of the 12 derived strains obtained after induction were classified as h-VISA according to the following criteria: area under the curve (AUC) on day 18/AUC of Mu3 of ≥90% and/or growth on brain heart infusion (BHI) agar with 4 mg/liter vancomycin. The derived isolates had thickened cell walls proportional to the level of glycopeptide resistance. Genes known to be associated with glycopeptide resistance (vraSR, yvqF, SA1703, graRS, walKR, and rpoB) were PCR sequenced; no de novo mutations were observed upon beta-lactam exposure. To determine whether trfA, a gene encoding a glycopeptide resistance factor, was essential in the selection of h-VISA upon beta-lactam pressure, a trfA-knockout strain was generated by allelic replacement. Indeed, beta-lactam exposure of this mutated strain showed no capacity to induce vancomycin resistance. In conclusion, these results showed that beta-lactam antibiotics at subinhibitory concentrations can induce intermediate vancomycin resistance in vitro. This induction required an intact trfA locus. Our results suggest that prior use of beta-lactam antibiotics can compromise vancomycin efficacy in the treatment of MRSA infections. PMID:24957836

  15. The effects of N-acylhomoserine lactones, β-lactam antibiotics and adenosine on biofilm formation in the multi-β-lactam antibiotic-resistant bacterium Acidovorax sp. strain MR-S7.

    PubMed

    Kusada, Hiroyuki; Hanada, Satoshi; Kamagata, Yoichi; Kimura, Nobutada

    2014-07-01

    Bacteria in the natural ecosystem frequently live as adherent communities called biofilms. Some chemical compounds are known to affect biofilm formation. We investigated the effect of exogenous small molecules, N-acylhomoserine lactones (AHLs), β-lactam antibiotics, and adenosine, on biofilm formation in the β-lactam antibiotic-resistant bacterium Acidovorax sp. strain MR-S7. Biofilm formation was induced by the addition of various types of AHL isomers and β-lactam antibiotics, whereas the addition of adenosine strongly interfered with the biofilm formation. A gene (macP) encoding adenosine deaminase (that converts adenosine to inosine controlling intracellular adenosine concentration) was successfully cloned from MR-S7 genome and heterologously expressed in Escherichia coli. The purified MacP protein clearly catalyzed the deamination of adenosine to produce inosine. A transcriptional analysis revealed that biofilm-inducing molecules, an AHL and a β-lactam antibiotic, strongly induced not only biofilm formation but also adenosine deaminase gene expression, suggesting that an elaborate gene regulation network for biofilm formation is present in the β-lactam antibiotic-resistant bacterium studied here.

  16. Profiling of β-Lactam Selectivity for Penicillin-Binding Proteins in Escherichia coli Strain DC2

    PubMed Central

    Kocaoglu, Ozden

    2015-01-01

    Penicillin-binding proteins (PBPs) are integral players in bacterial cell division, and their catalytic activities can be monitored with β-lactam-containing chemical probes. Compounds that target a single PBP could provide important information about the specific role(s) of each enzyme, making identification of such molecules important. We evaluated 22 commercially available β-lactams for inhibition of the PBPs in live Escherichia coli strain DC2. Whole cells were titrated with β-lactam antibiotics and subsequently incubated with a fluorescent penicillin derivative, Bocillin-FL (Boc-FL), to label uninhibited PBPs. Protein visualization was accomplished by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separation and fluorescent scanning. The examined β-lactams exhibited diverse PBP selectivities, with amdinocillin (mecillinam) showing selectivity for PBP2, aztreonam, piperacillin, cefuroxime, cefotaxime, and ceftriaxone for PBP3, and amoxicillin and cephalexin for PBP4. The remaining β-lactams did not block any PBPs in the DC2 strain of E. coli or inhibited more than one PBP at all examined concentrations in this Gram-negative organism. PMID:25733506

  17. Profiling of β-lactam selectivity for penicillin-binding proteins in Escherichia coli strain DC2.

    PubMed

    Kocaoglu, Ozden; Carlson, Erin E

    2015-05-01

    Penicillin-binding proteins (PBPs) are integral players in bacterial cell division, and their catalytic activities can be monitored with β-lactam-containing chemical probes. Compounds that target a single PBP could provide important information about the specific role(s) of each enzyme, making identification of such molecules important. We evaluated 22 commercially available β-lactams for inhibition of the PBPs in live Escherichia coli strain DC2. Whole cells were titrated with β-lactam antibiotics and subsequently incubated with a fluorescent penicillin derivative, Bocillin-FL (Boc-FL), to label uninhibited PBPs. Protein visualization was accomplished by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separation and fluorescent scanning. The examined β-lactams exhibited diverse PBP selectivities, with amdinocillin (mecillinam) showing selectivity for PBP2, aztreonam, piperacillin, cefuroxime, cefotaxime, and ceftriaxone for PBP3, and amoxicillin and cephalexin for PBP4. The remaining β-lactams did not block any PBPs in the DC2 strain of E. coli or inhibited more than one PBP at all examined concentrations in this Gram-negative organism. PMID:25733506

  18. Azide-alkyne cycloaddition en route towards 1H-1,2,3-triazole-tethered β-lactam-ferrocene and β-lactam-ferrocenylchalcone conjugates: synthesis and in vitro anti-tubercular evaluation.

    PubMed

    Kumar, Kewal; Carrère-Kremer, Séverine; Kremer, Laurent; Guérardel, Yann; Biot, Christophe; Kumar, Vipan

    2013-02-01

    A diverse range of triazoles were prepared following well established, Cu-mediated azide-alkyne cycloaddition reactions with the aim of probing the anti-tubercular structure-activity relationships (SAR) within the β-lactam-ferrocene-triazole conjugate family. The anti-tubercular evaluation studies of the synthesized conjugates revealed that none of the scaffolds exhibited any activity that restricted mycobacterial growth even at high doses. The introduction of various substituents onto the N-1 of the β-lactam ring, introducing mono- or di-ferrocenylchalcone substituents at the C-3 position as well as introducing a spacer of varying chain length failed to produce any significant enhancement in the activity profiles. The described protocol was a successful attempt on the inclusion of a ferrocene nucleus in the β-lactam family tethered via triazole linkers having metabolic stability and physicochemical favourability.

  19. Elaboration of a consensual definition of de-escalation allowing a ranking of β-lactams.

    PubMed

    Weiss, E; Zahar, J-R; Lesprit, P; Ruppe, E; Leone, M; Chastre, J; Lucet, J-C; Paugam-Burtz, C; Brun-Buisson, C; Timsit, J-F

    2015-07-01

    Empirical broad spectrum antimicrobial therapy prescribed in life-threatening situations should be de-escalated to mitigate the risk of resistance emergence. Definitions of de-escalation (DE) vary among studies, thereby biasing their results. The aim of this study was to provide a consensus definition of DE and to establish a ranking of β-lactam according to both their spectra and their ecological consequences. Twenty-eight experts from intensive care, infectious disease and clinical microbiology were consulted using the Delphi method (four successive questionnaires) from July to November 2013. More than 70% of similar answers to a question were necessary to reach a consensus. According to our consensus definition, DE purpose was to reduce both the spectrum of antimicrobial therapy and the selective pressure on microbiota. DE included switching from combination to monotherapy. A six-rank consensual classification of β-lactams allowing gradation of DE was established. The group was unable to differentiate ecological consequences of molecules included in group 4, i.e. piperacillin/tazobactam, ticarcillin/clavulanic acid, fourth-generation cephalosporin and antipseudomonal third-generation cephalosporin. Furthermore, no consensus was reached on the delay within which DE should be performed and on whether or not the shortening of antibiotic therapy duration should be included in DE definition. This study provides a consensual ranking of β-lactams according to their global ecological consequences that may be helpful in future studies on DE. However, this work also underlines the difficulties of reaching a consensus on the relative ecological impact of each individual drug and on the timing of DE.

  20. Profiling of β-Lactam Selectivity for Penicillin-Binding Proteins in Streptococcus pneumoniae D39

    PubMed Central

    Kocaoglu, Ozden; Tsui, Ho-Ching T.; Winkler, Malcolm E.

    2015-01-01

    Selective fluorescent β-lactam chemical probes enable the visualization of the transpeptidase activity of penicillin-binding proteins (PBPs) at different stages of bacterial cell division. To facilitate the development of new fluorescent probes for PBP imaging, we evaluated 20 commercially available β-lactams for selective PBP inhibition in an unencapsulated derivative of the D39 strain of Streptococcus pneumoniae. Live cells were treated with β-lactam antibiotics at different concentrations and subsequently incubated with Bocillin FL (Boc-FL; fluorescent penicillin) to saturate uninhibited PBPs. Fluorophore-labeled PBPs were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and fluorescence scanning. Among 20 compounds tested, carbapenems (doripenem and meropenem) were coselective for PBP1a, PBP2x, and PBP3, while six of the nine penicillin compounds were coselective for PBP2x and PBP3. In contrast, the seven cephalosporin compounds tested display variability in their PBP-binding profiles. Three cephalosporin compounds (cefoxitin, cephalexin, and cefsulodin) and the monobactam aztreonam exhibited selectivity for PBP3, while only cefuroxime (a cephalosporin) was selective for PBP2x. Treatment of S. pneumoniae cultures with a sublethal concentration of cefuroxime that inhibited 60% of PBP2x activity and less than 20% of the activity of other PBPs resulted in formation of elongated cells. In contrast, treatment of S. pneumoniae cultures with concentrations of aztreonam and cefoxitin that inhibited up to 70% of PBP3 activity and less than 30% of other PBPs resulted in no discernible morphological changes. Additionally, correlation of the MIC and IC50s for each PBP, with the exception of faropenem, amdinocillin (mecillinam), and 6-APA, suggests that pneumococcal growth inhibition is primarily due to the inhibition of PBP2x. PMID:25845878

  1. Antibacterial activity of RU44790, a new N-tetrazolyl monocyclic beta-lactam.

    PubMed Central

    Chantot, J F; Klich, M; Teutsch, G; Bryskier, A; Collette, P; Markus, A; Seibert, G

    1992-01-01

    RU44790 belongs to a new class of synthetic monocyclic beta-lactam antibiotics which feature a bioisosteric tetrazole moiety instead of the more classical acidic functions at the N-1 position of the beta-lactam ring. Its antibacterial activity was evaluated against some 900 strains and was compared with those of other recent beta-lactam derivatives, especially aztreonam. RU44790 is endowed with potent activity against gram-negative bacteria. At less than or equal to 0.6 micrograms/ml, RU44790 inhibited 90% of all strains of the family Enterobacteriaceae with the exception of Citrobacter spp. (MIC for 90% of strains tested, 1.2 micrograms/ml). The activity was similar to that of aztreonam against strains that are normally susceptible to expanded-spectrum cephalosporins. On the other hand, the new compound was 10 to 100 times more potent than aztreonam and most of the other antibiotics tested against enterobacteria that produce chromosome-encoded or plasmid-mediated extended-spectrum beta-lactamases. Pseudomonas aeruginosa isolates were equally susceptible to both monobactams. RU44790 was inactive against staphylococci and had only marginal activity against streptococci (MIC for 50% of strains tested, 2.5 micrograms/ml). RU44790 was highly resistant to hydrolysis by various beta-lactamases, particularly cephalosporinases such as P99. The latter enzyme was also inhibited by the compound. RU44790 showed a high affinity for penicillin-binding protein 3 of Escherichia coli. The results suggest that RU44790 has good potential in the treatment of infections caused by gram-negative microorganisms. Images PMID:1416860

  2. Profiling of β-lactam selectivity for penicillin-binding proteins in Streptococcus pneumoniae D39.

    PubMed

    Kocaoglu, Ozden; Tsui, Ho-Ching T; Winkler, Malcolm E; Carlson, Erin E

    2015-01-01

    Selective fluorescent β-lactam chemical probes enable the visualization of the transpeptidase activity of penicillin-binding proteins (PBPs) at different stages of bacterial cell division. To facilitate the development of new fluorescent probes for PBP imaging, we evaluated 20 commercially available β-lactams for selective PBP inhibition in an unencapsulated derivative of the D39 strain of Streptococcus pneumoniae. Live cells were treated with β-lactam antibiotics at different concentrations and subsequently incubated with Bocillin FL (Boc-FL; fluorescent penicillin) to saturate uninhibited PBPs. Fluorophore-labeled PBPs were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and fluorescence scanning. Among 20 compounds tested, carbapenems (doripenem and meropenem) were coselective for PBP1a, PBP2x, and PBP3, while six of the nine penicillin compounds were coselective for PBP2x and PBP3. In contrast, the seven cephalosporin compounds tested display variability in their PBP-binding profiles. Three cephalosporin compounds (cefoxitin, cephalexin, and cefsulodin) and the monobactam aztreonam exhibited selectivity for PBP3, while only cefuroxime (a cephalosporin) was selective for PBP2x. Treatment of S. pneumoniae cultures with a sublethal concentration of cefuroxime that inhibited 60% of PBP2x activity and less than 20% of the activity of other PBPs resulted in formation of elongated cells. In contrast, treatment of S. pneumoniae cultures with concentrations of aztreonam and cefoxitin that inhibited up to 70% of PBP3 activity and less than 30% of other PBPs resulted in no discernible morphological changes. Additionally, correlation of the MIC and IC50s for each PBP, with the exception of faropenem, amdinocillin (mecillinam), and 6-APA, suggests that pneumococcal growth inhibition is primarily due to the inhibition of PBP2x. PMID:25845878

  3. The influence of protein binding upon tissue fluid levels of six beta-lactam antibiotics.

    PubMed

    Wise, R; Gillett, A P; Cadge, B; Durham, S R; Baker, S

    1980-07-01

    The effect of protein binding upon the penetration of six-beta-lactam (three penicillins and three cephalosporins) antibiotics into tissue fluid was studied in humans. A cantharides blister technique was used. It was found that there was a linear relationship between the percentage of protein binding and the penetration into the blister fluid of the antibiotic as measured by the area under the curve of the protein-free fraction. This finding is further evidence that protein binding may have important influence upon the likely efficacy of an antimicrobial agent.

  4. Synthesis, antiviral activity, and bioavailability studies of gamma-lactam derived HIV protease inhibitors.

    PubMed

    Hungate, R W; Chen, J L; Starbuck, K E; Vacca, J P; McDaniel, S L; Levin, R B; Dorsey, B D; Guare, J P; Holloway, M K; Whitter, W

    1994-09-01

    Incorporation of a gamma-lactam in hydroxyethylene isosteres results in modest inhibitors of HIV-1 protease. Additional structural activity studies have produced significantly more potent inhibitors with the introduction of the trisubstituted cyclopentane (see compound 20) as the optimum substituent for the C-terminus. This new amino acid amide surrogate can be readily prepared in large scale from (R)-pulegone. Optimized compounds (36) and (60) are potent antiviral agents and are well absorbed (15-20%) in a dog model after oral administration. PMID:7712123

  5. Alterations in peptidoglycan of Neisseria gonorrhoeae induced by sub-MICs of beta-lactam antibiotics.

    PubMed Central

    Garcia-Bustos, J F; Dougherty, T J

    1987-01-01

    Exposure of Neisseria gonorrhoeae to sub-MICs of selected beta-lactam antibiotics caused distortion of normal cell morphology. Analysis of the peptidoglycan indicated that the cells were accumulating increased quantities of disaccharide pentapeptide in their cell walls. The O-acetylated form of the disaccharide pentapeptide was not detected among the major peaks. The correlation of antibiotic binding to gonococcal penicillin-binding protein 2 and accumulation of non-O-acetylated disaccharide pentapeptide suggested an explanation for the previously observed relationship of penicillin-binding protein 2 and O-acetylation of peptidoglycan. PMID:3105447

  6. Microwave-assisted solid-phase synthesis of side-chain to side-chain lactam-bridge cyclic peptides.

    PubMed

    Tala, Srinivasa R; Schnell, Sathya M; Haskell-Luevano, Carrie

    2015-12-15

    Side-chain to side-chain lactam-bridged cyclic peptides have been utilized as therapeutic agents and biochemical tools. Previous synthetic methods of these peptides need special reaction conditions, form side products and take longer reaction times. Herein, an efficient microwave-assisted synthesis of side-chain to side-chain lactam-bridge cyclic peptides SHU9119 and MTII is reported. The synthesis time and efforts are significantly reduced in the present method, without side product formation. The analytical and pharmacological data of the synthesized cyclic peptides are in accordance with the commercially obtained compounds. This new method could be used to synthesize other side-chain to side-chain lactam-bridge peptides and amenable to automation and extensive SAR compound derivatization.

  7. Protein Binding of β-Lactam Antibiotics in Critically Ill Patients: Can We Successfully Predict Unbound Concentrations?

    PubMed Central

    Wong, Gloria; Briscoe, Scott; Adnan, Syamhanin; McWhinney, Brett; Ungerer, Jacobus; Lipman, Jeffrey

    2013-01-01

    The use of therapeutic drug monitoring (TDM) to optimize beta-lactam dosing in critically ill patients is growing in popularity, although there are limited data describing the potential impact of altered protein binding on achievement of target concentrations. The aim of this study was to compare the measured unbound concentration to the unbound concentration predicted from published protein binding values for seven beta-lactams using data from blood samples obtained from critically ill patients. From 161 eligible patients, we obtained 228 and 220 plasma samples at the midpoint of the dosing interval and trough, respectively, for ceftriaxone, cefazolin, meropenem, piperacillin, ampicillin, benzylpenicillin, and flucloxacillin. The total and unbound beta-lactam concentrations were measured using validated methods. Variabilities in both unbound and total concentrations were marked for all antibiotics, with significant differences being present between measured and predicted unbound concentrations for ceftriaxone and for flucloxacillin at the mid-dosing interval (P < 0.05). The predictive performance for calculating unbound concentrations using published protein binding values was poor, with bias for overprediction of unbound concentrations for ceftriaxone (83.3%), flucloxacillin (56.8%), and benzylpenicillin (25%) and underprediction for meropenem (12.1%). Linear correlations between the measured total and unbound concentrations were observed for all beta-lactams (R2 = 0.81 to 1.00; P < 0.05) except ceftriaxone and flucloxacillin. The percent protein binding of flucloxacillin and the plasma albumin concentration were also found to be linearly correlated (R2 = 0.776; P < 0.01). In conclusion, significant differences between measured and predicted unbound drug concentrations were found only for the highly protein-bound beta-lactams ceftriaxone and flucloxacillin. However, direct measurement of unbound drug in research and clinical practice is suggested for selected

  8. Identification of a structural determinant for resistance to β-lactam antibiotics in Gram-positive bacteria

    PubMed Central

    Mouz, N.; Gordon, E.; Di Guilmi, A.-M.; Petit, I.; Pétillot, Y.; Dupont, Y.; Hakenbeck, R.; Vernet, T.; Dideberg, O.

    1998-01-01

    Streptococcus pneumoniae is the main causal agent of pathologies that are increasingly resistant to antibiotic treatment. Clinical resistance of S. pneumoniae to β-lactam antibiotics is linked to multiple mutations of high molecular mass penicillin-binding proteins (H-PBPs), essential enzymes involved in the final steps of bacterial cell wall synthesis. H-PBPs from resistant bacteria have a reduced affinity for β-lactam and a decreased hydrolytic activity on substrate analogues. In S. pneumoniae, the gene coding for one of these H-PBPs, PBP2x, is located in the cell division cluster (DCW). We present here structural evidence linking multiple β-lactam resistance to amino acid substitutions in PBP2x within a buried cavity near the catalytic site that contains a structural water molecule. Site-directed mutation of amino acids in contact with this water molecule in the “sensitive” form of PBP2x produces mutants similar, in terms of β-lactam affinity and substrate hydrolysis, to altered PBP2x produced in resistant clinical isolates. A reverse mutation in a PBP2x variant from a clinically important resistant clone increases the acylation efficiency for β-lactams and substrate analogues. Furthermore, amino acid residues in contact with the structural water molecule are conserved in the equivalent H-PBPs of pathogenic Gram-positive cocci. We suggest that, probably via a local structural modification, the partial or complete loss of this water molecule reduces the acylation efficiency of PBP2x substrates to a point at which cell wall synthesis still occurs, but the sensitivity to therapeutic concentrations of β-lactam antibiotics is lost. PMID:9811812

  9. Important Role of a Putative Lytic Transglycosylase Cj0843c in β-Lactam Resistance in Campylobacter jejuni

    PubMed Central

    Zeng, Ximin; Gillespie, Barbara; Lin, Jun

    2015-01-01

    Beta-lactam antibiotics are an important class of antibiotics for treating bacterial infections. Despite prevalent β-lactam resistance in Campylobacter jejuni, the leading bacterial cause of human diarrhea in developed countries, molecular mechanism of β-lactam resistance in C. jejuni is still largely unknown. In this study, C. jejuni 81–176 was used for random transposon mutagenesis. Screening of a 2,800-mutant library identified 22 mutants with increased susceptibility to ampicillin. Of these mutants, two mutants contains mutations in Cj0843c (a putative lytic transglycosylase gene) and in its upstream gene Cj0844c, respectively. Complementation experiment demonstrated that the Cj0843 contributes to β-lactam resistance. The Cj0843c insertional mutation was subsequently introduced to diverse C. jejuni clinical strains for MIC test, showing that Cj0843c contributes to both intrinsic and acquired β-lactam resistance of C. jejuni. Consistent with this finding, inactivation of Cj0843c also dramatically reduced β-lactamase activity. Genomic examination and PCR analysis showed Cj0843c is widely distributed in C. jejuni. High purity recombinant Cj0843c was produced for generation of specific antiserum. The Cj0843 was localized in the periplasm, as demonstrated by immunoblotting using specific antibodies. Turbidimetric assay further demonstrated the capability of the purified Cj0843c to hydrolyze cell walls. Inactivation of Cj0843c also significantly reduced C. jejuni colonization in the intestine. Together, this study identifies a mechanism of β-lactam resistance in C. jejuni and provides insights into the role of cell wall metabolism in regulating β-lactamase activity. PMID:26635760

  10. Clickable 4-Oxo-β-lactam-Based Selective Probing for Human Neutrophil Elastase Related Proteomes.

    PubMed

    Ruivo, Eduardo F P; Gonçalves, Lídia M; Carvalho, Luís A R; Guedes, Rita C; Hofbauer, Stefan; Brito, José A; Archer, Margarida; Moreira, Rui; Lucas, Susana D

    2016-09-20

    Human neutrophil elastase (HNE) is a serine protease associated with several inflammatory processes such as chronic obstructive pulmonary disease (COPD). The precise involvement of HNE in COPD and other inflammatory disease mechanisms has yet to be clarified. Herein we report a copper-catalyzed alkyne-azide 1,3-dipolar cycloaddition (CuAAC, or 'click' chemistry) approach based on the 4-oxo-β-lactam warhead that yielded potent HNE inhibitors containing a triazole moiety. The resulting structure-activity relationships set the basis to develop fluorescent and biotinylated activity-based probes as tools for molecular functional analysis. Attaching the tags to the 4-oxo-β-lactam scaffold did not affect HNE inhibitory activity, as revealed by the IC50 values in the nanomolar range (56-118 nm) displayed by the probes. The nitrobenzoxadiazole (NBD)-based probe presented the best binding properties (ligand efficiency (LE)=0.31) combined with an excellent lipophilic ligand efficiency (LLE=4.7). Moreover, the probes showed adequate fluorescence properties, internalization in human neutrophils, and suitable detection of HNE in the presence of a large excess of cell lysate proteins. This allows the development of activity-based probes with promising applications in target validation and identification, as well as diagnostic tools. PMID:27465595

  11. Rapid cytolysis of Mycobacterium tuberculosis by faropenem, an orally bioavailable β-lactam antibiotic.

    PubMed

    Dhar, Neeraj; Dubée, Vincent; Ballell, Lluis; Cuinet, Guillaume; Hugonnet, Jean-Emmanuel; Signorino-Gelo, François; Barros, David; Arthur, Michel; McKinney, John D

    2015-02-01

    Recent clinical studies indicate that meropenem, a β-lactam antibiotic, is a promising candidate for therapy of drug-resistant tuberculosis. However, meropenem is chemically unstable, requires frequent intravenous injection, and must be combined with a β-lactamase inhibitor (clavulanate) for optimal activity. Here, we report that faropenem, a stable and orally bioavailable β-lactam, efficiently kills Mycobacterium tuberculosis even in the absence of clavulanate. The target enzymes, L,D-transpeptidases, were inactivated 6- to 22-fold more efficiently by faropenem than by meropenem. Using a real-time assay based on quantitative time-lapse microscopy and microfluidics, we demonstrate the superiority of faropenem to the frontline antituberculosis drug isoniazid in its ability to induce the rapid cytolysis of single cells. Faropenem also showed superior activity against a cryptic subpopulation of nongrowing but metabolically active cells, which may correspond to the viable but nonculturable forms believed to be responsible for relapses following prolonged chemotherapy. These results identify faropenem to be a potential candidate for alternative therapy of drug-resistant tuberculosis. PMID:25421469

  12. Rapid Cytolysis of Mycobacterium tuberculosis by Faropenem, an Orally Bioavailable β-Lactam Antibiotic

    PubMed Central

    Dhar, Neeraj; Dubée, Vincent; Ballell, Lluis; Cuinet, Guillaume; Hugonnet, Jean-Emmanuel; Signorino-Gelo, François; Arthur, Michel; McKinney, John D.

    2014-01-01

    Recent clinical studies indicate that meropenem, a β-lactam antibiotic, is a promising candidate for therapy of drug-resistant tuberculosis. However, meropenem is chemically unstable, requires frequent intravenous injection, and must be combined with a β-lactamase inhibitor (clavulanate) for optimal activity. Here, we report that faropenem, a stable and orally bioavailable β-lactam, efficiently kills Mycobacterium tuberculosis even in the absence of clavulanate. The target enzymes, l,d-transpeptidases, were inactivated 6- to 22-fold more efficiently by faropenem than by meropenem. Using a real-time assay based on quantitative time-lapse microscopy and microfluidics, we demonstrate the superiority of faropenem to the frontline antituberculosis drug isoniazid in its ability to induce the rapid cytolysis of single cells. Faropenem also showed superior activity against a cryptic subpopulation of nongrowing but metabolically active cells, which may correspond to the viable but nonculturable forms believed to be responsible for relapses following prolonged chemotherapy. These results identify faropenem to be a potential candidate for alternative therapy of drug-resistant tuberculosis. PMID:25421469

  13. A new tetracyclic lactam building block for thick, broad-bandgap photovoltaics.

    PubMed

    Kroon, Renee; Diaz de Zerio Mendaza, Amaia; Himmelberger, Scott; Bergqvist, Jonas; Bäcke, Olof; Faria, Gregório Couto; Gao, Feng; Obaid, Abdulmalik; Zhuang, Wenliu; Gedefaw, Desta; Olsson, Eva; Inganäs, Olle; Salleo, Alberto; Müller, Christian; Andersson, Mats R

    2014-08-20

    A new tetracyclic lactam building block for polymer semiconductors is reported that was designed to combine the many favorable properties that larger fused and/or amide-containing building blocks can induce, including improved solid-state packing, high charge carrier mobility, and improved charge separation. Copolymerization with thiophene resulted in a semicrystalline conjugated polymer, PTNT, with a broad bandgap of 2.2 eV. Grazing incidence wide-angle X-ray scattering of PTNT thin films revealed a strong tendency for face-on π-stacking of the polymer backbone, which was retained in PTNT:fullerene blends. Corresponding solar cells featured a high open-circuit voltage of 0.9 V, a fill factor around 0.6, and a power conversion efficiency as high as 5% for >200 nm thick active layers, regardless of variations in blend stoichiometry and nanostructure. Moreover, efficiencies of >4% could be retained when thick active layers of ∼400 nm were employed. Overall, these values are the highest reported for a conjugated polymer with such a broad bandgap and are unprecedented in materials for tandem and particularly ternary blend photovoltaics. Hence, the newly developed tetracyclic lactam unit has significant potential as a conjugated building block in future organic electronic materials. PMID:25056482

  14. Alpha-melanocyte-stimulating hormone suppresses oxidative stress through a p53-mediated signaling pathway in human melanocytes.

    PubMed

    Kadekaro, Ana Luisa; Chen, Juping; Yang, Jennifer; Chen, Shuna; Jameson, Joshua; Swope, Viki B; Cheng, Tan; Kadakia, Madhavi; Abdel-Malek, Zalfa

    2012-06-01

    Epidermal melanocytes are skin cells specialized in melanin production. Activation of the melanocortin 1 receptor (MC1R) on melanocytes by α-melanocyte-stimulating hormone (α-MSH) induces synthesis of the brown/black pigment eumelanin that confers photoprotection from solar UV radiation (UVR). Contrary to keratinocytes, melanocytes are slow proliferating cells that persist in the skin for decades, in an environment with high levels of UVR-induced reactive oxygen species (ROS). We previously reported that in addition to its role in pigmentation, α-MSH also reduces oxidative stress and enhances the repair of DNA photoproducts in melanocytes, independent of melanin synthesis. Given the significance of ROS in carcinogenesis, here we investigated the mechanisms by which α-MSH exerts antioxidant effects in melanocytes. We show that activation of the MC1R by α-MSH contributes to phosphorylation of p53 on serine 15, a known requirement for stabilization and activation of p53, a major sensor of DNA damage. This effect is mediated by the cAMP/PKA pathway and by the activation of phosphoinositide 3-kinase (PI3K) ATR and DNA protein kinase (DNA-PK). α-MSH increases the levels of 8-oxoguanine DNA glycosylase (OGG1) and apurinic apyrimidinic endonuclease 1 (APE-1/Ref-1), enzymes essential for base excision repair. Nutlin-3, an HDM2 inhibitor, mimicked the effects of α-MSH resulting in reduced phosphorylation of H2AX (γ-H2AX), a marker of DNA damage. Conversely, the p53 inhibitor pifithrin-α or silencing of p53 abolished the effects of α-MSH and augmented oxidative stress. These results show that p53 is an important target of the downstream MC1R signaling that reduces oxidative stress and possibly malignant transformation of melanocytes.

  15. alpha-Melanocyte stimulating hormone (MSH) decreases cyclosporine a induced apoptosis in cultured human proximal tubular cells.

    PubMed Central

    Jo, S. K.; Lee, S. Y.; Han, S. Y.; Cha, D. R.; Cho, W. Y.; Kim, H. K.; Won, N. H.

    2001-01-01

    The pathogenesis of chronic cyclosporine A (CsA) nephrotoxicity has not been elucidated, but apoptosis is thought to play an important role in CsA induced tubular atrophy. Recently Fas-Fas ligand system mediated apoptosis has been frequently reported in many epithelial cells as well as in T lymphocytes. We investigated the ability of CsA to induce apoptosis in cultured human proximal tubular epithelial cells and also the effect of alpha-MSH on them. Fas, Fas ligand, and an intracellular adaptor protein, Fas-associating protein with death domain (FADD) expression, and poly-ADP ribose polymerase (PARP) cleavage were also studied. CsA induced apoptosis in cultured tubular epithelial cells demonstrated by increased number of TUNEL positive cells and it was accompanied by a significant increase in Fas mRNA and Fas ligand protein expressions. FADD and the cleavage product of PARP also increased, indicating the activation of caspase. In alpha-MSH co-treated cells, apoptosis markedly decreased with downregulation of Fas, Fas ligand and FADD expressions and also the cleavage product of PARP. In conclusion, these data suggest that tubular cell apoptosis mediated by Fas system may play a role in tubular atrophy in chronic CsA nephrotoxicity and pretreatment of alpha-MSH may have a some inhibitory effect on CsA induced tubular cell apoptosis. PMID:11641530

  16. TarO-specific inhibitors of wall teichoic acid biosynthesis restore β-lactam efficacy against methicillin-resistant staphylococci.

    PubMed

    Lee, Sang Ho; Wang, Hao; Labroli, Marc; Koseoglu, Sandra; Zuck, Paul; Mayhood, Todd; Gill, Charles; Mann, Paul; Sher, Xinwei; Ha, Sookhee; Yang, Shu-Wei; Mandal, Mihir; Yang, Christine; Liang, Lianzhu; Tan, Zheng; Tawa, Paul; Hou, Yan; Kuvelkar, Reshma; DeVito, Kristine; Wen, Xiujuan; Xiao, Jing; Batchlett, Michelle; Balibar, Carl J; Liu, Jenny; Xiao, Jianying; Murgolo, Nicholas; Garlisi, Charles G; Sheth, Payal R; Flattery, Amy; Su, Jing; Tan, Christopher; Roemer, Terry

    2016-03-01

    The widespread emergence of methicillin-resistant Staphylococcus aureus (MRSA) has dramatically eroded the efficacy of current β-lactam antibiotics and created an urgent need for new treatment options. We report an S. aureus phenotypic screening strategy involving chemical suppression of the growth inhibitory consequences of depleting late-stage wall teichoic acid biosynthesis. This enabled us to identify early-stage pathway-specific inhibitors of wall teichoic acid biosynthesis predicted to be chemically synergistic with β-lactams. We demonstrated by genetic and biochemical means that each of the new chemical series discovered, herein named tarocin A and tarocin B, inhibited the first step in wall teichoic acid biosynthesis (TarO). Tarocins do not have intrinsic bioactivity but rather demonstrated potent bactericidal synergy in combination with broad-spectrum β-lactam antibiotics against diverse clinical isolates of methicillin-resistant staphylococci as well as robust efficacy in a murine infection model of MRSA. Tarocins and other inhibitors of wall teichoic acid biosynthesis may provide a rational strategy to develop Gram-positive bactericidal β-lactam combination agents active against methicillin-resistant staphylococci.

  17. A fluorescent carbapenem for structure function studies of penicillin-binding proteins, β-lactamases, and β-lactam sensors.

    PubMed

    June, Cynthia M; Vaughan, Robert M; Ulberg, Lucas S; Bonomo, Robert A; Witucki, Laurie A; Leonard, David A

    2014-10-15

    By reacting fluorescein isothiocyanate with meropenem, we have prepared a carbapenem-based fluorescent β-lactam. Fluorescein-meropenem binds both penicillin-binding proteins and β-lactam sensors and undergoes a typical acylation reaction in the active site of these proteins. The probe binds the class D carbapenemase OXA-24/40 with close to the same affinity as meropenem and undergoes a complete catalytic hydrolysis reaction. The visible light excitation and strong emission of fluorescein render this molecule a useful structure-function probe through its application in sodium dodecyl sulfate-polyacrylamide gel electrophoresis assays as well as solution-based kinetic anisotropy assays. Its classification as a carbapenem β-lactam and the position of its fluorescent modification render it a useful complement to other fluorescent β-lactams, most notably Bocillin FL. In this study, we show the utility of fluorescein-meropenem by using it to detect mutants of OXA-24/40 that arrest at the acyl-intermediate state with carbapenem substrates but maintain catalytic competency with penicillin substrates.

  18. Emergence of cross-resistance to imipenem and other beta-lactam antibiotics in Pseudomonas aeruginosa during therapy.

    PubMed

    Pagani, L; Landini, P; Luzzaro, F; Debiaggi, M; Romero, E

    1990-01-01

    The emergence of resistance to imipenem and other beta-lactams by Pseudomonas aeruginosa was investigated with two pairs of isolates. Two of these isolates were susceptible to imipenem and other beta-lactam antibiotics, such as moxalactam, ceftriaxone and cefotaxime, while the other two had developed resistance to those antibiotics during imipenem therapy. So far imipenem-resistant isolates have not demonstrated cross-resistance to other beta-lactam agents. We examined in these clinical isolates the possible mechanisms of resistance due to permeability modifications, either in outer membrane proteins (porins) or to LPS (lipopolysaccharides) complex. Particularly we analysed possible modification of physico-chemical properties of outer membrane proteins, such as changes in their hydrophobicity and electrical charge. beta-lactamase production was also studied. Results showed that resistance to imipenem may be related to loss or modifications in hydrophobicity of an outer membrane protein of about 46 Kdal; other modifications concerned hydrophobicity of the porin OMP F and, in one strain, the LPS complex appears to be responsible for resistance to other beta-lactam antibiotics together in combination with the production of beta-lactamases.

  19. TarO-specific inhibitors of wall teichoic acid biosynthesis restore β-lactam efficacy against methicillin-resistant staphylococci.

    PubMed

    Lee, Sang Ho; Wang, Hao; Labroli, Marc; Koseoglu, Sandra; Zuck, Paul; Mayhood, Todd; Gill, Charles; Mann, Paul; Sher, Xinwei; Ha, Sookhee; Yang, Shu-Wei; Mandal, Mihir; Yang, Christine; Liang, Lianzhu; Tan, Zheng; Tawa, Paul; Hou, Yan; Kuvelkar, Reshma; DeVito, Kristine; Wen, Xiujuan; Xiao, Jing; Batchlett, Michelle; Balibar, Carl J; Liu, Jenny; Xiao, Jianying; Murgolo, Nicholas; Garlisi, Charles G; Sheth, Payal R; Flattery, Amy; Su, Jing; Tan, Christopher; Roemer, Terry

    2016-03-01

    The widespread emergence of methicillin-resistant Staphylococcus aureus (MRSA) has dramatically eroded the efficacy of current β-lactam antibiotics and created an urgent need for new treatment options. We report an S. aureus phenotypic screening strategy involving chemical suppression of the growth inhibitory consequences of depleting late-stage wall teichoic acid biosynthesis. This enabled us to identify early-stage pathway-specific inhibitors of wall teichoic acid biosynthesis predicted to be chemically synergistic with β-lactams. We demonstrated by genetic and biochemical means that each of the new chemical series discovered, herein named tarocin A and tarocin B, inhibited the first step in wall teichoic acid biosynthesis (TarO). Tarocins do not have intrinsic bioactivity but rather demonstrated potent bactericidal synergy in combination with broad-spectrum β-lactam antibiotics against diverse clinical isolates of methicillin-resistant staphylococci as well as robust efficacy in a murine infection model of MRSA. Tarocins and other inhibitors of wall teichoic acid biosynthesis may provide a rational strategy to develop Gram-positive bactericidal β-lactam combination agents active against methicillin-resistant staphylococci. PMID:26962156

  20. Copper-catalyzed intermolecular dehydrogenative amidation/amination of quinoline N-oxides with lactams/cyclamines.

    PubMed

    Li, Gang; Jia, Chunqi; Sun, Kai

    2013-10-18

    C-H, N-H dehydrogenative coupling of quinoline N-oxides with lactams/cyclamines has been achieved in the presence of the Cu(OAc)2 catalyst to give good to excellent yields. This study provides a new strategy for the construction of a 2-aminoquinoline skeleton via direct functionalization of aryl C-H bonds. PMID:24093556

  1. In vitro activities of 15 oral beta-lactams against Klebsiella pneumoniae harboring new extended-spectrum beta-lactamases.

    PubMed Central

    Kitzis, M D; Liassine, N; Ferré, B; Gutmann, L; Acar, J F; Goldstein, F

    1990-01-01

    The activities of 15 oral beta-lactams against Klebsiella pneumoniae harboring new extended-spectrum beta-lactamases were studied. All compounds were affected by these enzymes, especially by the SHV derivatives. Except for ceftibuten, the compounds with the greatest intrinsic activity were more affected by the presence of these enzymes than were older compounds with moderate intrinsic activity. PMID:2285291

  2. [Isolation of the ergot (Claviceps purpurea (Fr.) Tul., strain VKM-F-366D), producing the lactamic alkaloid ergocornam].

    PubMed

    Komarova, E L; Shain, S S; Sheĭchenko, V I

    2002-01-01

    A new ergot strain VKM-F-3662D producing lactamic alkaloid ergocornam with concomitant alkaloids valinamide and ergometrine was isolated during selective works with sclerotium MS-462, which was obtained from ergocryptine ergot strain VKM-F-2642D. The structure of these alkaloids was determined by 1H and 13C NMR. PMID:12449796

  3. Reactivation of peptidoglycan synthesis in ether-permeabilized Escherichia coli after inhibition by beta-lactam antibiotics.

    PubMed Central

    Talbot, M K; Schaefer, F; Brocks, V; Christenson, J G

    1989-01-01

    The recovery of peptidoglycan-synthesizing activity after inhibition by beta-lactam antibiotics was investigated in ether-permeabilized cells of Escherichia coli B. Such cells synthesize sodium dodecyl sulfate-insoluble peptidoglycan when provided with UDP-linked precursors and Mg2+. The ability of beta-lactam antibiotics to inhibit the synthesis of peptidoglycan was correlated with their affinity for penicillin-binding proteins 1A and 1Bs. Penicillin-binding protein 1Bs is thought to be the major peptidoglycan synthetase in E. coli and is a major lethal target for beta-lactam antibiotics. Ether-treated bacteria were preincubated with concentrations of beta-lactams sufficient to completely inhibit peptidoglycan synthesis and then treated with beta-lactamases to inactivate free antibiotic prior to measurement of peptidoglycan synthesis. At 40 min after beta-lactamase treatment, the rate of peptidoglycan synthesis was about 74% of the control rate in cells pretreated with ampicillin, but only 15% of the control in cells pretreated with penicillin G or azlocillin. Reversal of inhibition by several other antibiotics fell between these extremes. When cross-linking of peptidoglycan was measured specifically, reversal of inhibition by ampicillin also occurred more readily than that by penicillin G. Reactivation of peptidoglycan synthesis was not due to de novo synthesis of penicillin-binding proteins since it occurred under conditions that did not allow incorporation of [14C]leucine. We conclude that there is considerable variation in the stability of the inactive acyl enzymes formed between various beta-lactams and penicillin-binding protein 1Bs, with those formed by penicillin G being relatively long-lived. Images PMID:2515794

  4. An HPLC-DAD method for the simultaneous determination of nine β-lactam antibiotics in ewe milk.

    PubMed

    Cámara, M; Gallego-Picó, A; Garcinuño, R M; Fernández-Hernando, P; Durand-Alegría, J S; Sánchez, P J

    2013-11-15

    The presence of β-lactam residues in foodstuffs constitutes a potential risk to the human health and undesirable effects on consumers, and nowadays these antibiotic residues are also recognised as an emerging environmental problem. In addition, these are of great concern to prestigious Manchego cheese processors (Central Spain denomination of origin) because they reduce the curdling of milk and cause improper cheese ripening, which consequently lead to an important loss of monetary income. This work describes the development of a sensitive and reliable method using liquid chromatography with UV-diode array detection (LC-DAD) for simultaneous determination of the β-lactam antibiotics, ampicillin (AMP), benzylpenicillin (PEG), cephalexin (CFX), cefazolin (CFL), cefoperazone (CFP), cloxacillin (CLO), dicloxacillin (DCL), oxacillin (OXA) and phenoxymethylpenicillin (PEV), in Manchega ewe milk. The column, mobile phase, temperature and flow rate were optimised to provide the best resolution of these analytes. The extraction method of the antibiotic residues involves the deproteinisation of the milk sample using acetonitrile and centrifugation followed by a solid-phase extraction (SPE) clean-up. The recoveries for the studied β-lactams ranged from 79% to 96% with relative standard deviations between 0.5% and 4.9%. The limits of quantification (LOQs) for all these compounds were in the range of 3.4-8.6μgkg(-1), which are lower than the maximum residue limits (MRLs) established by the European Union for the studied β-lactams in milk, making the method suitable for performing routine analyses. The proposed multi-residue LC-UV-diode array detection (LC-DAD) method is a powerful and popular alternative for the determination and confirmation of antibiotic residues in small milk industries and is the first one capable of determining nine β-lactam antibiotics in samples of Manchega ewe milk. PMID:23790854

  5. Contribution of peptidoglycan amidation to beta-lactam and lysozyme resistance in different genetic lineages of Staphylococcus aureus.

    PubMed

    Figueiredo, Teresa A; Ludovice, Ana Madalena; Sobral, Rita G

    2014-06-01

    The enzymes responsible for peptidoglycan amidation in Staphylococcus aureus, MurT and GatD, were recently identified and shown to be required for optimal expression of resistance to beta-lactams, bacterial growth, and resistance to lysozyme. In this study, we analyzed the impact of peptidoglycan amidation in representative strains of the most widespread clones of methicillin resistant S. aureus (MRSA). The inhibition of the expression of murT-gatD operon resulted in different phenotypes of resistance to beta-lactams and lysozyme according to the different genetic backgrounds. Further, clonal lineages CC1 and CC398 (community-acquired MRSA [CA-MRSA]) showed a stronger dependency on MurT-GatD for resistance to beta-lactams, when compared to the impact of the impairment of the cell wall step catalyzed by MurF. In the remaining backgrounds similar phenotypes of beta-lactam resistance were observed upon the impairment of both cell-wall-related genes. Therefore, for CA-related backgrounds, the predominant beta-lactam resistance mechanism seems to involve genes associated with secondary modifications of peptidoglycan. On the other hand, the lack of glutamic acid amidation had a more substantial impact on lysozyme resistance for cells of CA-MRSA backgrounds, than for hospital-acquired MRSA (HA-MRSA). However, no significant differences were found in the resistance level of the respective peptidoglycan structure, suggesting that the lysozyme resistance mechanism involves other factors. Taken together, these results suggested that the different genetic lineages of MRSA were able to develop different molecular strategies to overcome the selective pressures experienced during evolution.

  6. Lactam nonanic acid, a new substance from Cleome viscosa with allelopathic and antimicrobial properties.

    PubMed

    Jana, Anirban; Biswas, Suparna Mandal

    2011-03-01

    Cleome viscosa L. (Capparidaceae) is well known for its medicinal properties. Lactam nonanoic acid (LNA) [2-amino-9-(4-oxoazetidin-2-yl)-nonanoic acid; C12H22N2O3, mol. wt. 242] has been isolated and purified from the root exudates of Cleome viscosa. The aqueous solution of this pure compound has been tested on bacteria (Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus) and fungi (Aspergillus fumigatus, A. niger and A. tamarii). At a dosage of 500 ppm and above, P. aeruginosa and S. aureus were totally inhibited while E. coli remained unaffected. On the other hand, growth of A. niger and A. tamarii was stimulated while there was no effect on A. fumigatus. This pure compound showed concentration-dependent inhibitory activity on rice, gram and mustard seeds. PMID:21451245

  7. Polymers bearing groups derived from n-substituted lactams and their use as lubricating oil additives

    SciTech Connect

    Brulet, D.; Chauvel, B.; Pocheville, R.

    1980-09-16

    Novel lubricating oil polymer additives are obtained by the following: (1) by preparing, by anionic polymerization, a living diene polymer of mn of between about 20,000 and 300,000; (2) by functionalizing the said polymer by means of an nsubstituted lactam of the type of n-alkylcaprolactam, nbinylcaprolactam, and particularly of the type of nalkylpyrrolidione and n-vinylpyrrolidone; and (3) by hydrogenating the said functionalized polymer. A variant method of preparing the said polymers comprises subjecting the living polymer to a metalation operation before functionalization; the hydrogenation operation is carried out before metalation or after functionalization. The said polymers may be used as additives which improve the viscosity index and the dispersing power of lubricating oils. The amount of additive added is between about 0.1 and 10 percent by weight.

  8. Regulation of the vitamin D receptor by vitamin D lactam derivatives.

    PubMed

    Asano, Lisa; Waku, Tsuyoshi; Abe, Rumi; Kuwabara, Naoyuki; Ito, Ichiaki; Yanagisawa, Junn; Nagasawa, Kazuo; Shimizu, Toshiyuki

    2016-09-01

    The active metabolite of vitamin D3 , 1α,25-dihydroxyvitamin D3 , acts as a ligand for the vitamin D receptor (VDR) and activates VDR-mediated gene expression. Recently, we characterized 1α,25-dihydroxyvitamin D3 -26,23-lactams (DLAMs), which mimic vitamin D3 metabolites, as noncalcemic VDR ligands that barely activate the receptor. In this study, we present structural insights onto the regulation of VDR function by DLAMs. X-ray crystallographic analysis revealed that DLAMs induced a large conformational change in the loop region between helices H6 and H7 in the VDR ligand-binding domain. Our structural analysis suggests that targeting of the loop region may be a new mode of VDR regulation. PMID:27500498

  9. Crystal structure of human renal dipeptidase involved in beta-lactam hydrolysis.

    PubMed

    Nitanai, Yasushi; Satow, Yoshinori; Adachi, Hideki; Tsujimoto, Masafumi

    2002-08-01

    Human renal dipeptidase is a membrane-bound glycoprotein hydrolyzing dipeptides and is involved in hydrolytic metabolism of penem and carbapenem beta-lactam antibiotics. The crystal structures of the saccharide-trimmed enzyme are determined as unliganded and inhibitor-liganded forms. They are informative for designing new antibiotics that are not hydrolyzed by this enzyme. The active site in each of the (alpha/beta)(8) barrel subunits of the homodimeric molecule is composed of binuclear zinc ions bridged by the Glu125 side-chain located at the bottom of the barrel, and it faces toward the microvillar membrane of a kidney tubule. A dipeptidyl moiety of the therapeutically used cilastatin inhibitor is fully accommodated in the active-site pocket, which is small enough for precise recognition of dipeptide substrates. The barrel and active-site architectures utilizing catalytic metal ions exhibit unexpected similarities to those of the murine adenosine deaminase and the catalytic domain of the bacterial urease.

  10. Cooperative Catalysis by Carbenes and Lewis Acids in a Highly Stereoselective Route to γ-Lactams

    PubMed Central

    Raup, Dustin E. A.; Cardinal-David, Benoit; Holte, Dane; Scheidt, Karl A.

    2010-01-01

    Enzymes are a continuing source of inspiration for the design of new chemical reactions that proceed with efficiency, high selectivity and minimal waste. In many biochemical processes, different catalytic species, such as Lewis acids and bases, are involved in precisely orchestrated interactions to activate reactants simultaneously or sequentially. Employing this type of cooperative catalysis, in which two or more catalytic cycles operate concurrently to achieve one overall transformation, has great potential to enhance known reactivity and drive the development of new chemical reactions with high value. In this disclosure, a cooperative N-heterocyclic carbene/Lewis acid catalytic system promotes the addition of homoenolate equivalents to hydrazones generating highly substituted γ-lactams in moderate to good yields and high levels of diastereo- and enantioselectivity. PMID:20729898

  11. Modularity and three-dimensional isostructurality of novel synthons in sulfonamide–lactam cocrystals

    PubMed Central

    Bolla, Geetha; Mittapalli, Sudhir; Nangia, Ashwini

    2015-01-01

    The design of novel supramolecular synthons for functional groups relevant to drugs is an essential prerequisite for applying crystal engineering in the development of novel pharmaceutical cocrystals. It has been convincingly shown over the past decade that molecular level control and modulation can influence the physicochemical properties of drug cocrystals. Whereas considerable advances have been reported on the design of cocrystals for carboxylic acids and carboxamide functional groups, the sulfonamide group, which is a cornerstone of sulfa drugs, is relatively unexplored for reproducible heterosynthon-directed crystal engineering. The occurrence of synthons and isostructurality in sulfonamide–lactam cocrystals (SO2NH2⋯CONH hydrogen bonding) is analyzed to define a strategy for amide-type GRAS (generally recognized as safe) coformers with sulfonamides. Three types of supramolecular synthons are identified for the N—H donor of sulfonamide hydrogen bonding to the C=O acceptor of amide. Synthon 1: catemer synthon C 2 1(4) chain motif, synthon 2: dimer–cyclic ring synthon R 2 2(8)R 4 2(8) motifs, and synthon 3: dimer–catemer synthon of R 2 2(8)C 1 1(4)D notation. These heterosynthons of the cocrystals observed in this study are compared with the N—H⋯O dimer R 2 2(8) ring and C(4) chain motifs of the individual sulfonamide structures. The X-ray crystal structures of sulfonamide–lactam cocrystals exhibit interesting isostructurality trends with the same synthon being present. One-dimensional, two-dimensional and three-dimensional isostructurality in crystal structures is associated with isosynthons and due to their recurrence, novel heterosynthons for sulfonamide cocrystals are added to the crystal engineer’s toolkit. With the predominance of sulfa drugs in medicine, these new synthons provide rational strategies for the design of binary and potentially ternary cocrystals of sulfonamides. PMID:26175899

  12. Quantitative assessment of faecal shedding of β-lactam-resistant Escherichia coli and enterococci in dogs.

    PubMed

    Espinosa-Gongora, Carmen; Shah, Syed Qaswar Ali; Jessen, Lisbeth Rem; Bortolaia, Valeria; Langebæk, Rikke; Bjørnvad, Charlotte Reinhard; Guardabassi, Luca

    2015-12-31

    Quantitative data on faecal shedding of antimicrobial resistant bacteria are crucial to assess the risk of transmission from dogs to other animals as well as humans. In this study we investigated prevalence and concentrations of β-lactam-resistant Escherichia coli and enterococci in the faeces of 108 dogs presenting at a veterinary hospital in Denmark. The dogs had not been treated with antimicrobials for 4 weeks prior to the study. Total E. coli and enterococci were quantified by counts on MacConkey and Slanetz-Bartley, respectively. Resistant E. coli and enterococci were counted on the same media containing relevant antibiotic concentrations, followed by species identification using MALDI-TOF. Ampicillin- and cefotaxime-resistant E. coli were detected in 40% and 8% of the dogs, respectively, whereas approximately 15% carried ampicillin-resistant enterococci, mainly Enterococcus faecium. In the faeces of the carriers, the proportion of resistant strains in the total bacterial species population was on average 15% for both ampicillin-resistant E. coli (median faecal load 3.2×10(4)cfu/g) and E. faecium (5.8×10(2) cfu/g), and 4.6% for cefotaxime-resistant E. coli (8.6×10(3) cfu/g). Cefotaxime resistance was associated with the presence of blaCTX-M-1 (n=4), blaCMY-2 (n=4) or multiple mutations in the promoter and coding region of chromosomal ampC (n=1). Altogether the results indicate that the risks of zoonotic transmission of β-lactam-resistant bacteria via human exposure to canine faeces greatly vary amongst individual dogs and are influenced by unidentified factors other than recent antimicrobial use.

  13. Recognition and clinical significance of mechanisms of bacterial resistance to beta-lactams.

    PubMed

    Mouton, R P

    1984-01-01

    Resistance to beta-lactams may be difficult to recognize. This is due to the difficulty in detecting these resistances, when the routine tests performed in diagnostic laboratories are interpreted in the usual manner. Since failure to recognize this type of resistance may have serious consequences for the patient, it is essential that it be detected when present. For the detection of methicillin resistance of Staphylococcus aureus a standardized method using either a medium containing 5% NaCl or a low incubation temperature is advocated. Methicillin resistance of S. epidermidis can only be recognized reliably by means of a quantitative test and incubation for 42-48 h. Resistance of Haemophilus influenzae to ampicillin may be intrinsic or it may be caused by a TEM beta-lactamase; a beta-lactamase test should be used to detect the latter type of resistance. Inducible cephalosporinase may be responsible for the rapid development of resistance of some bacterial species to cefamandole, even during therapy. If a stable beta-lactamase production is attained by mutation, resistance to other beta-lactams will usually be present as well. Routine induction tests should be performed for all isolates of species of Enterobacter, Serratia, Citrobacter and Proteus, indole-positive. The same type of 'hidden' resistance may be present in Pseudomonas aeruginosa, with regard to cefotaxime and other third-generation cephalosporins. Beta-lactamase-positive Neisseria gonorrhoeae can easily be recognized by a beta-lactamase test. In addition, the results of diffusion tests allow one to distinguish between beta-lactamase-positive and beta-lactamase-negative strains. Recognition of those strains of N. gonorrhoeae having a decreased susceptibility to penicillin is only possible when well-standardized quantitative tests are used.

  14. Persistence and degradation of new β-lactam antibiotics in the soil and water environment.

    PubMed

    Braschi, I; Blasioli, S; Fellet, C; Lorenzini, R; Garelli, A; Pori, M; Giacomini, D

    2013-09-01

    The development of new antibiotics with low environmental persistence is of utmost importance in contrasting phenomena of antibiotic resistance. In this study, the persistence of two newly synthesized monocyclic β-lactam antibiotics: (2R)-1-(methylthio)-4-oxoazetidin-2-yl acetate, P1, and (2R,3R)-3-((1R)-1-(tert-butyldimethylsilanyloxy)ethyl)-1-(methylthio)-4-oxoazetidin-2-yl acetate, P2, has been investigated in water in the pH range 3-9 and in two (calcareous and forest) soils, then compared to amoxicillin, a β-lactam antibiotic used in human and veterinary medicine. P1 and P2 persistence in water was lower than that of amoxicillin with only a few exceptions. P1 hydrolysis was catalyzed at an acidic pH whereas P2 hydrolysis takes place at both acidic and alkaline pH values. P1 persistence in soils depended mainly on their water potential (t1/2: 35.0-70.7d at wilting point; <1d at field capacity) whereas for P2 it was shorter and unaffected by soil water content (t1/2 0.13-2.5d). Several degradation products were detected in soils at both water potentials, deriving partly from hydrolytic pathways and partly from microbial transformation. The higher LogKow value for P2 compared with P1 seemingly confers P2 with high permeability to microbial membranes regardless of soil water content. P1 and P2 persistence in soils at wilting point was shorter than that of amoxicillin, whereas it had the same extent at field capacity.

  15. Metal-based biologically active azoles and β-lactams derived from sulfa drugs.

    PubMed

    Ebrahimi, Hossein Pasha; Hadi, Jabbar S; Almayah, Abdulelah A; Bolandnazar, Zeinab; Swadi, Ali G; Ebrahimi, Amirpasha

    2016-03-01

    Metal complexes of Schiff bases derived from sulfamethoxazole (SMZ) and sulfathiazole (STZ), converted to their β-lactam derivatives have been synthesized and experimentally characterized by elemental analysis, spectral (IR, (1)H NMR, (13)C NMR, and EI-mass), molar conductance measurements and thermal analysis techniques. The structural and electronic properties of the studied molecules were investigated theoretically by performing density functional theory (DFT) to access reliable results to the experimental values. The spectral and thermal analysis reveals that the Schiff bases act as bidentate ligands via the coordination of azomethine nitrogen to metal ions as well as the proton displacement from the phenolic group through the metal ions; therefore, Cu complexes can attain the square planner arrangement and Zn complexes have a distorted tetrahedral structure. The thermogravimetric (TG/DTG) analyses confirm high stability for all complexes followed by thermal decomposition in different steps. In addition, the antibacterial activities of synthesized compounds have been screened in vitro against various pathogenic bacterial species. Inspection of the results revealed that all newly synthesized complexes individually exhibit varying degrees of inhibitory effects on the growth of the tested bacterial species, therefore, they may be considered as drug candidates for bacterial pathogens. The free Schiff base ligands (1-2) exhibited a broad spectrum antibacterial activity against Gram negative Escherichia coli, Pseudomonas aeruginosa, and Proteus spp., and Gram positive Staphylococcus aureus bacterial strains. The results also indicated that the β-lactam derivatives (3-4) have high antibacterial activities on Gram positive bacteria as well as the metal complexes (5-8), particularly Zn complexes, have a significant activity against all Gram negative bacterial strains. It has been shown that the metal complexes have significantly higher activity than corresponding

  16. Recognition and clinical significance of mechanisms of bacterial resistance to beta-lactams.

    PubMed

    Mouton, R P

    1984-01-01

    Resistance to beta-lactams may be difficult to recognize. This is due to the difficulty in detecting these resistances, when the routine tests performed in diagnostic laboratories are interpreted in the usual manner. Since failure to recognize this type of resistance may have serious consequences for the patient, it is essential that it be detected when present. For the detection of methicillin resistance of Staphylococcus aureus a standardized method using either a medium containing 5% NaCl or a low incubation temperature is advocated. Methicillin resistance of S. epidermidis can only be recognized reliably by means of a quantitative test and incubation for 42-48 h. Resistance of Haemophilus influenzae to ampicillin may be intrinsic or it may be caused by a TEM beta-lactamase; a beta-lactamase test should be used to detect the latter type of resistance. Inducible cephalosporinase may be responsible for the rapid development of resistance of some bacterial species to cefamandole, even during therapy. If a stable beta-lactamase production is attained by mutation, resistance to other beta-lactams will usually be present as well. Routine induction tests should be performed for all isolates of species of Enterobacter, Serratia, Citrobacter and Proteus, indole-positive. The same type of 'hidden' resistance may be present in Pseudomonas aeruginosa, with regard to cefotaxime and other third-generation cephalosporins. Beta-lactamase-positive Neisseria gonorrhoeae can easily be recognized by a beta-lactamase test. In addition, the results of diffusion tests allow one to distinguish between beta-lactamase-positive and beta-lactamase-negative strains. Recognition of those strains of N. gonorrhoeae having a decreased susceptibility to penicillin is only possible when well-standardized quantitative tests are used. PMID:6442123

  17. Investigation of β-lactam antibacterial drugs, β-lactamases, and penicillin-binding proteins with fluorescence polarization and anisotropy: a review

    NASA Astrophysics Data System (ADS)

    Shapiro, Adam B.

    2016-06-01

    This review covers the uses of fluorescence polarization and anisotropy for the investigation of bacterial penicillin binding proteins (PBPs), which are the targets of β-lactam antibacterial drugs (penicillins, cephalosporins, carbapenems, and monobactams), and of the β-lactamase enzymes that destroy these drugs and help to render bacterial pathogens resistant to them. Fluorescence polarization and anisotropy-based methods for quantitation of β-lactam drugs are also reviewed. A particular emphasis is on methods for quantitative measurement of the interactions of β-lactams and other inhibitors with PBPs and β-lactamases.

  18. Reductive Amination/Cyclization of Keto Acids Using a Hydrosilane for Selective Production of Lactams versus Cyclic Amines by Switching of the Indium Catalyst.

    PubMed

    Ogiwara, Yohei; Uchiyama, Takuya; Sakai, Norio

    2016-01-26

    Described herein is that the catalytic construction of N-substituted five- and six-membered lactams from keto acids with primary amines by reductive amination, using an indium/silane combination. This relatively benign and safe catalyst/reductant system tolerates the use of a variety of functional groups, especially ones that are reduction-sensitive. A direct switch from synthesizing lactams to synthesizing cyclic amines is achieved by changing the catalyst from In(OAc)3 to InI3. This conversion occurs by further reduction of the lactam using the indium/silane pair.

  19. In silico analysis of different generation β lactams antibiotics with penicillin binding protein-2 of Neisseria meningitidis for curing meningococcal disease.

    PubMed

    Tripathi, Vijay; Tripathi, Pooja; Srivastava, Navita; Gupta, Dwijendra

    2014-12-01

    Neisseria meningitidis is a gram negative, diplococcic pathogen responsible for the meningococcal disease and fulminant septicemia. Penicillin-binding proteins-2 (PBPs) is crucial for the cell wall biosynthesis during cell proliferation of N. meningitidis and these are the target for β-lactam antibiotics. For many years penicillin has been recognized as the antibiotic for meningococcal disease but the meningococcus has seemed to be antibiotic resistance. In the present work we have verified the molecular interaction of Penicillin binding protein-2 N. meningitidis to different generation of β-lactam antibiotics and concluded that the third generation of β-lactam antibiotics shows efficient binding with Penicillin binding protein-2 of N. meningitidis. On the basis of binding efficiency and inhibition constant, ceftazidime emerged as the most efficient antibiotic amongst the other advanced β-lactam antibiotics against Penicillin-binding protein-2 of N. meningitidis.

  20. Structure-activity analysis of the growth hormone secretagogue GHRP-6 by alpha- and beta-amino gamma-lactam positional scanning.

    PubMed

    Boutard, Nicolas; Jamieson, Andrew G; Ong, Huy; Lubell, William D

    2010-01-01

    Incorporation of amino lactams into biologically active peptides restricts conformational mobility and may enhance selectivity and increase potency. alpha- and beta-amino gamma-lactams (Agl and Bgl), in both S and R configurations, were introduced into the growth hormone secretagogue GHRP-6 using a Fmoc-compatible solid-phase protocol relying on N-alkylation with five- and six-membered cyclic sulfamidates, followed by lactam annulation under microwave heating. Using this protocol in conjunction with IRORI Kan techniques furnished eleven new GHRP-6 analogs, and their binding affinity IC50 values on both the growth hormone secretagogue receptor 1a (GHS-R1a) and CD36 receptors are herein reported. The results indicate that selectivity towards one receptor or the other can be modulated by lactam substitution, typically at the Ala3 and the D-Phe5 positions. PMID:19954433

  1. Synthesis of Strained γ-Lactams by Palladium(0)-Catalyzed C(sp(3) )-H Alkenylation and Application to Alkaloid Synthesis.

    PubMed

    Holstein, Philipp M; Dailler, David; Vantourout, Julien; Shaya, Janah; Millet, Anthony; Baudoin, Olivier

    2016-02-18

    A variety of strained α-alkylidene-γ-lactams were synthesized by palladium(0)-catalyzed intramolecular C(sp(3) )-H alkenylation from easily accessible acyclic and monocyclic bromoalkene precursors. These lactams are valuable intermediates for accessing various classes of mono- and bicylic alkaloids containing a pyrrolidine ring, as illustrated with the synthesis of an advanced model of the marine natural product plakoridine A and of the indolizidine alkaloid δ-coniceine.

  2. Structurally novel Bi- and tricyclic beta-lactams via [2 + 2] cycloaddition or radical reactions in 2-azetidinone-tethered enallenes and allenynes.

    PubMed

    Alcaide, Benito; Almendros, Pedro; Aragoncillo, Cristina

    2003-10-16

    [reaction: see text] Thermolysis of beta-lactam-tethered enallenyl alcohols gave tricyclic ring structures via a formal [2 + 2] cycloaddition of the alkene with the distal bond of the allene, while the tin-promoted radical cyclization in 2-azetidinone-tethered allenynes proceeded to provide bicyclic beta-lactams containing a medium-sized ring. The access to cyclization precursors was achieved by regio- and stereoselective metal-mediated carbonyl allenylation of 4-oxoazetidine-2-carbaldehydes in an aqueous environment.

  3. In vitro susceptibility of Campylobacter jejuni to 27 antimicrobial agents and various combinations of beta-lactams with clavulanic acid or sulbactam.

    PubMed Central

    Van der Auwera, P; Scorneaux, B

    1985-01-01

    The in vitro susceptibility of human isolates of Campylobacter jejuni was investigated with 27 antibiotics and 8 combinations of beta-lactams with clavulanic acid or sulbactam. Ansamycin, the new quinolines, erythromycin, and cefpirome were the most active drugs against C. jejuni; amoxicillin, ampicillin, cefotaxime, and ceftazidime 90% of the isolates, greater than or equal to 50 mg/liter). The activity of various beta-lactams was unchanged by the addition of clavulanic acid or sulbactam. PMID:2994557

  4. Regioselective Synthesis of a Family of β‐Lactams Bearing a Triazole Moiety as Potential Apoptosis Inhibitors

    PubMed Central

    Garrido, Maria; Corredor, Miriam; Orzáez, Mar; Alfonso, Ignacio

    2016-01-01

    Abstract Apoptosis is a biological process important to several human diseases; it is strongly regulated through protein–protein interactions and complex formation. We previously reported the synthesis of apoptosis inhibitors bearing an exocyclic triazole amide isoster by using an Ugi four‐component coupling reaction (Ugi‐4CC), followed by a base‐promoted intramolecular cyclization. Depending on the substitution patterns and the reaction conditions, this cyclization forms the six‐ or four‐membered ring. Two compounds bearing the β‐lactam scaffold turned out to be the most potent inhibitors. This encouraged us to optimize the modulation of the cyclization, and prepare a library of 15 β‐lactams with total regioselectivity. Moreover, we aimed to improve the bioavailability of these compounds through the introduction of diversity at different substitution positions. The activity of these compounds as apoptosis inhibitors in cellular extracts has been evaluated, showing an increase in their potency. PMID:27777842

  5. Are new antibiotics better than beta-lactams for non-critical inpatients with community-acquired pneumonia?

    PubMed

    Reyes B, Tomás; Ortega G, Marcos; Saldías P, Fernando

    2016-01-01

    Treatment for community-acquired pneumonia in immunocompetent adults is mainly empirical. Beta-lactam antibiotics have been traditionally considered first-line therapy. New antibiotics could be more effective but the evidence is not clear until now, and its use could entail greater costs, an increase in bacterial resistance and other adverse effects. Searching in Epistemonikos database, which is maintained by screening 30 databases, we identified six systematic reviews including 36 randomized trials addressing this question. We combined the evidence using meta-analysis and generated a summary of findings table following the GRADE approach. We concluded new antibiotics are not better than beta-lactam antibiotics for the treatment of non-critical inpatients with community-acquired pneumonia in relation to clinical failure or adverse effects. PMID:27512983

  6. [Cross allergy between penicillins and other beta lactam antibiotics--the risk is much less than previously thought].

    PubMed

    Tängden, Thomas; Furebring, Mia; Löwdin, Elisabeth; Werner, Sonja

    2015-02-03

    Severe IgE-mediated allergic reactions to penicillins are rare but might be fatal. Because some studies demonstrated a high risk of cross-sensitivity to cephalosporins and carbapenems it has been recommended to avoid these antibiotics in patients with suspected hypersensitivity to penicillins. However, recent studies and analyses conclude that the risk of cross-reactivity was overestimated in the earlier studies and that it is in fact very low for parenteral cephalosporins and perhaps even negligible for carbapenems. The new knowledge has implications for the choice of therapy for bacterial infections in patients with a history of penicillin hypersensitivity, because alternative antibiotic regimens are often inferior to beta-lactam antibiotics. The aim of the present review is to present existing knowledge on cross-sensitivity between beta-lactams, as well as to discuss the management of patients with suspected allergic reactions to these antibiotics.

  7. [In vitro inhibition of granulopoiesis by beta-lactam antibiotics. Comparison of piperacillin, mezlocillin, ceftriaxone and ceftazidime].

    PubMed

    Marie, J P; Thevenin, D; Zittoun, R

    1986-12-20

    The mechanism of neutropenia induced by beta-lactam antibiotics was explored by studying the action of these drugs on granulopoiesis in vitro. Normal bone marrows were cultivated in the presence of increasing concentrations of piperacillin (10 marrows), mezlocillin, ceftriaxone and ceftazidime (5 marrows each) in order to find out whether these antibiotics exhibited toxicity to granulocyte-monocyte precursors. A dose-dependent inhibition of granulopoiesis was found in all cases. When the doses used were equivalent to maximum plasma concentrations in vivo, inhibition was minimal with piperacillin and mezlocillin and much more pronounced with the cephalosporins. This dose-dependent inhibition suggests that toxicity is involved in the mechanism of neutropenia induced by beta-lactam antibiotics.

  8. Are new antibiotics better than beta-lactams for non-critical inpatients with community-acquired pneumonia?

    PubMed

    Reyes B, Tomás; Ortega G, Marcos; Saldías P, Fernando

    2016-08-05

    Treatment for community-acquired pneumonia in immunocompetent adults is mainly empirical. Beta-lactam antibiotics have been traditionally considered first-line therapy. New antibiotics could be more effective but the evidence is not clear until now, and its use could entail greater costs, an increase in bacterial resistance and other adverse effects. Searching in Epistemonikos database, which is maintained by screening 30 databases, we identified six systematic reviews including 36 randomized trials addressing this question. We combined the evidence using meta-analysis and generated a summary of findings table following the GRADE approach. We concluded new antibiotics are not better than beta-lactam antibiotics for the treatment of non-critical inpatients with community-acquired pneumonia in relation to clinical failure or adverse effects.

  9. Sustainable Chiral Polyamides with High Melting Temperature via Enhanced Anionic Polymerization of a Menthone-Derived Lactam.

    PubMed

    Winnacker, Malte; Neumeier, Michael; Zhang, Xiaohan; Papadakis, Christine M; Rieger, Bernhard

    2016-05-01

    Polyamides are very important polymers that find applications from commodities up to the automotive and biomedical sectors, and their impact is continuously growing. The synthesis of structurally significant, chiral, and sustainable polyamides is described via a new, convenient, and solvent-free anionic polymerization of a biobased ε-lactam, which is obtained from the renewable terpenoid ketone l-menthone in a one-step synthesis. These polyamides are shown to have outstanding structural and thermal properties, which are thus introduced via the structure and chirality of the natural lactam monomer and which are discussed and compared with those of petroleum-based, established, and commercial polyamide Nylon-6. X-ray data reveal a remarkable degree of crystallinity in these green polymers and emphasize the impact of their structural features on the resulting properties. PMID:26992085

  10. Bioactive 7-Oxabicyclic[6.3.0]lactam and 12-Membered Macrolides from a Gorgonian-Derived Cladosporium sp. Fungus

    PubMed Central

    Cao, Fei; Yang, Qin; Shao, Chang-Lun; Kong, Chui-Jian; Zheng, Juan-Juan; Liu, Yun-Feng; Wang, Chang-Yun

    2015-01-01

    One new bicyclic lactam, cladosporilactam A (1), and six known 12-membered macrolides (2–7) were isolated from a gorgonian-derived Cladosporium sp. fungus collected from the South China Sea. Their complete structural assignments were elucidated by comprehensive spectroscopic investigation. Quantum chemistry calculations were used in support of the structural determination of 1. The absolute configuration of 1 was determined by calculation of its optical rotation. Cladosporilactam A (1) was the first example of 7-oxabicyclic[6.3.0]lactam obtained from a natural source. Compound 1 exhibited promising cytotoxic activity against cervical cancer HeLa cell line with an IC50 value of 0.76 μM. PMID:26198234

  11. Bioactive 7-Oxabicyclic[6.3.0]lactam and 12-Membered Macrolides from a Gorgonian-Derived Cladosporium sp. Fungus.

    PubMed

    Cao, Fei; Yang, Qin; Shao, Chang-Lun; Kong, Chui-Jian; Zheng, Juan-Juan; Liu, Yun-Feng; Wang, Chang-Yun

    2015-07-01

    One new bicyclic lactam, cladosporilactam A (1), and six known 12-membered macrolides (2-7) were isolated from a gorgonian-derived Cladosporium sp. fungus collected from the South China Sea. Their complete structural assignments were elucidated by comprehensive spectroscopic investigation. Quantum chemistry calculations were used in support of the structural determination of 1. The absolute configuration of 1 was determined by calculation of its optical rotation. Cladosporilactam A (1) was the first example of 7-oxabicyclic[6.3.0]lactam obtained from a natural source. Compound 1 exhibited promising cytotoxic activity against cervical cancer HeLa cell line with an IC50 value of 0.76 μM.

  12. Treatment of Gram-negative pneumonia in the critical care setting: is the beta-lactam antibiotic backbone broken beyond repair?

    PubMed

    Bassetti, Matteo; Welte, Tobias; Wunderink, Richard G

    2016-01-01

    Beta-lactam antibiotics form the backbone of treatment for Gram-negative pneumonia in mechanically ventilated patients in the intensive care unit. However, this beta-lactam antibiotic backbone is increasingly under pressure from emerging resistance across all geographical regions, and health-care professionals in many countries are rapidly running out of effective treatment options. Even in regions that currently have only low levels of resistance, the effects of globalization are likely to increase local pressures on the beta-lactam antibiotic backbone in the near future. Therefore, clinicians are increasingly faced with a difficult balancing act: the need to prescribe adequate and appropriate antibiotic therapy while reducing the emergence of resistance and the overuse of antibiotics. In this review, we explore the burden of Gram-negative pneumonia in the critical care setting and the pressure that antibiotic resistance places on current empiric therapy regimens (and the beta-lactam antibiotic backbone) in this patient population. New treatment approaches, such as systemic and inhaled antibiotic alternatives, are on the horizon and are likely to help tackle the rising levels of beta-lactam antibiotic resistance. In the meantime, it is imperative that the beta-lactam antibiotic backbone of currently available antibiotics be supported through stringent antibiotic stewardship programs.

  13. Treatment of Gram-negative pneumonia in the critical care setting: is the beta-lactam antibiotic backbone broken beyond repair?

    PubMed

    Bassetti, Matteo; Welte, Tobias; Wunderink, Richard G

    2016-01-01

    Beta-lactam antibiotics form the backbone of treatment for Gram-negative pneumonia in mechanically ventilated patients in the intensive care unit. However, this beta-lactam antibiotic backbone is increasingly under pressure from emerging resistance across all geographical regions, and health-care professionals in many countries are rapidly running out of effective treatment options. Even in regions that currently have only low levels of resistance, the effects of globalization are likely to increase local pressures on the beta-lactam antibiotic backbone in the near future. Therefore, clinicians are increasingly faced with a difficult balancing act: the need to prescribe adequate and appropriate antibiotic therapy while reducing the emergence of resistance and the overuse of antibiotics. In this review, we explore the burden of Gram-negative pneumonia in the critical care setting and the pressure that antibiotic resistance places on current empiric therapy regimens (and the beta-lactam antibiotic backbone) in this patient population. New treatment approaches, such as systemic and inhaled antibiotic alternatives, are on the horizon and are likely to help tackle the rising levels of beta-lactam antibiotic resistance. In the meantime, it is imperative that the beta-lactam antibiotic backbone of currently available antibiotics be supported through stringent antibiotic stewardship programs. PMID:26821535

  14. Increased susceptibility to beta-lactam antibiotics and decreased porin content caused by envB mutations of Salmonella typhimurium.

    PubMed Central

    Oppezzo, O J; Avanzati, B; Antón, D N

    1991-01-01

    Isogenic derivatives carrying envB6, envB9, or envB+ alleles were obtained from a strain of Salmonella typhimurium that was partially resistant to mecillinam, a beta-lactam antibiotic specific for penicillin-binding protein 2 (PBP 2). Testing of the isogenic strains with several antibacterial agents demonstrated that envB mutations either increased resistance (mecillinam) or did not affect the response (imipemen) to beta-lactams that act primarily on PBP 2, while susceptibilities to beta-lactams that act on PBP 1B, PBP 3, or both were increased. Furthermore, the susceptibilities of envB strains to hydrophobic compounds such as rifampin, novobiocin, or chloramphenicol were not modified, even though their susceptibilities to deoxycholate and crystal violet were enhanced. Outer cell membranes of envB mutants presented a 50% reduction in protein content compared with that of the isogenic envB+ strains, and OmpF and OmpD porins were particularly affected by the reduction. No alteration in the amount or pattern of periplasmic proteins was noticed, and lipopolysaccharides from envB mutants appeared to be normal by sodium dodecyl sulfate-urea-polyacrylamide gel electrophoresis. By using derivatives that produced a plasmid-encoded beta-lactamase, it was demonstrated that envB cells are slightly less permeable to cephalothin than envB+ bacteria are. It is concluded that the high susceptibility of envB mutants to beta-lactams is due to the increased effectiveness of the antibiotics on PBP 1B, PBP 3, or both. Images PMID:1656857

  15. Relation between Resistance to Antipseudomonal β-Lactams and ampC and mexC Genes of Pseudomonas aeruginosa

    PubMed Central

    Rezaei, Fatemeh; Saderi, Horieh; Boroumandi, Shahrsam; Faghihzadeh, Soghrat

    2016-01-01

    Background: In order to select a better antibiotic choice for treatment of Pseudomonas aeruginosa infections, this study was conducted to determine the frequency of resistance to some antipseudomonal β-lactams in P. aeruginosa isolates from patients in Tehran, Iran. In addition, the relation between presence of genes known to be responsible for resistance to β-lactams (ampC, mexC1,2, and mexC3,4 genes) and resistance phenotype among P. aeroginosa isolates was evaluated. Methods: P. aeruginosa strains were isolated and identified by routine methods and PCR for oprL gene. Disk diffusion method was employed to determine the antimicrobial susceptibility pattern according to CLSI recommendations. PCR was used to detect the resistance genes. Results: Among 100 isolates of P. aeruginosa, 82% had ampC, 86% mexC1,2 and 89% mexC3,4 genes and combinations of these genes were seen in most of isolates and only 3% of isolates had none of these genes. Resistance to mezlocillin, cefepime, ceftazidime and piperacillin/ tazobactam was seen in 46%, 41%, 36% and 29% of isolates, respectively. Significant relation (P value ≤0.05 by Chi-square or Fisher Exact test) was observed between the presence of ampC gene and resistance to all the studied β-lactams in this study. No relation was observed for mexC genes, although many of isolates containing these two genes were phenotypically resistant. Discussion: This study had shown for the first time, the presence of ampC and mexC genes in significant percent of clinical isolates of P. aeruginosa in Tehran, Iran, and relation between presence of ampC gene and resistance to β-lactams. PMID:26870143

  16. Effect of clavulanic acid on susceptibility of Campylobacter jejuni and Campylobacter coli to eight beta-lactam antibiotics.

    PubMed Central

    Gaudreau, C L; Lariviere, L A; Lauzer, J C; Turgeon, F F

    1987-01-01

    The effect of clavulanic acid on the susceptibility of 32 strains of Campylobacter jejuni and Campylobacter coli to eight beta-lactam agents was studied. Almost all strains tested became susceptible to amoxicillin and ticarcillin with 1 microgram of clavulanic acid per ml. This compound had little or no effect on susceptibility to penicillin G, cephalothin, cefamandole, and cefoxitin. Clavulanic acid had a marginal effect on cefotaxime and moxalactam susceptibility. PMID:3619428

  17. ELECTROSPRAY IONIZATION (ESI) FRAGMENTATIONS AND DIMETHYLDIOXIRANE REACTIVITIES OF THREE DIVERSE LACTAMS HAVING FULL, HALF, AND ZERO RESONANCE ENERGIES

    PubMed Central

    Morgan, Kathleen M.; Ashline, David J.; Morgan, Jessica P.; Greenberg, Arthur

    2014-01-01

    Three lactams having respectively ~20 kcal/mol, ~10 kcal/mol, and 0 kcal/mol of resonance energy have been subjected to electrospray ionization mass spectrometry (ESI/MS) as well as to attempted reaction with dimethyldioxirane (DMDO). The ESI/MS for all three lactams are consistent with fragmentation from the N-protonated, rather than the O-protonated tautomer. Each exhibits a unique fragmentation pathway. DFT calculations are employed to provide insights concerning these pathways. N-Ethyl-2-pyrrolidinone and 1-azabicyclo[3.3.1]nonan-2-one, the full- and half-resonance lactams, are unreactive with DMDO. The “Kirby lactam” (3,5,7-trimethyl-1-azaadamantan-2-one), has zero resonance energy and reacts rapidly with DMDO to generate a mixture of reaction products. The structure assigned to one of these is the 2,2-dihydroxy-N-oxide, thought to be stabilized by intramolecular hydrogen bonding and buttressing by the methyl substituents. A reasonable pathway to this derivative might involve formation of an extremely labile N-oxide, in a purely formal sense an example of the hithertounknown amide N-oxides, followed by hydration with traces of moisture. PMID:24313276

  18. Metallo-beta-lactamases of Pseudomonas aeruginosa--a novel mechanism resistance to beta-lactam antibiotics.

    PubMed

    Sacha, Paweł; Wieczorek, Piotr; Hauschild, Tomasz; Zórawski, Marcin; Olszańska, Dorota; Tryniszewska, Elzbieta

    2008-01-01

    Since about twenty years, following the introduction into therapeutic of news beta-lactam antibiotics (broad-spectrum cephalosporins, monobactams and carbapenems), a very significant number of new beta-lactamases appeared. These enzymes confer to the bacteria which put them, the means of resisting new molecules. The genetic events involved in this evolution are of two types: evolution of old enzymes by mutation and especially appearance of new genes coming for some, from bacteria of the environment. Numerous mechanisms of enzymatic resistance to the carbapenems have been described in Pseudomonas aeruginosa. The important mechanism of inactivation carbapenems is production variety of b-lactam hydrolysing enzymes associated to carbapenemases. The metallo-beta-enzymes (IMP, VIM, SPM, GIM types) are the most clinically significant carbapenemases. P. aeruginosa posses MBLs and seem to have acquired them through transmissible genetic elements (plasmids or transposons associated with integron) and can be transmission to other bacteria. They have reported worldwide but mostly from South East Asia and Europe. The enzymes, belonging to the molecular class B family, are the most worrisome of all beta-lactamases because they confer resistance to carbapenems and all the beta-lactams (with the exception of aztreonam) and usually to aminoglycosides and quinolones. The dissemination of MBLs genes is thought to be driven by regional consumption of extended--spectrum antibiotics (e.g. cephalosporins and carbapenems), and therefore care must be taken that these drugs are not used unnecessarily.

  19. Resistance to oxyimino beta-lactams due to a mutation of chromosomal beta-lactamase in Citrobacter freundii.

    PubMed

    Haruta, S; Nukaga, M; Taniguchi, K; Sawai, T

    1998-01-01

    The duplicative mutation of an Ala-Val-Arg sequence at positions 208 to 210 in the loop structure of Enterobacter cloacae class C beta-lactamase caused substrate specificity extension to oxyimino beta-lactam antibiotics and this chromosomal mutation provided bacterial cells with high resistance to the beta-lactams (M. Nukaga et al, 1995, J. Biol. Chem. 270, 5729-5735). In order to confirm the universality of this phenomenon among other class C beta-lactamases, the duplicative mutation was applied to a class C beta-lactamase of Citrobacter freundii, which has 74% homology to the E. cloacae beta-lactamase amino acid sequence. The counterpart sequence to the Ala-Val-Arg of the E. cloacae enzyme in C. freundii beta-lactamase was identified to be Pro-Val-His. A Pro-Val-His sequence was inserted just after the native Pro-Val-His sequence at positions 208 to 210 in the C. freundii beta-lactamase. The resulting mutant of C. freundii beta-lactamase obtained a striking characteristic that we expected, showing substrate specificity extension to oxyimino beta-lactams. Nearly the same result was obtained with the insertion of an Ala-Val-Arg sequence after the native Pro-Val-His sequence. These results indicate that structural modification of this locus commonly induces modification of the substrate specificity to unfavorable substrates for many chromosomal class C beta-lactamases produced by gram-negative bacteria.

  20. Factors essential for L,D-transpeptidase-mediated peptidoglycan cross-linking and β-lactam resistance in Escherichia coli

    PubMed Central

    Hugonnet, Jean-Emmanuel; Mengin-Lecreulx, Dominique; Monton, Alejandro; den Blaauwen, Tanneke; Carbonnelle, Etienne; Veckerlé, Carole; Brun, Yves, V.; van Nieuwenhze, Michael; Bouchier, Christiane; Tu, Kuyek; Rice, Louis B; Arthur, Michel

    2016-01-01

    The target of β-lactam antibiotics is the D,D-transpeptidase activity of penicillin-binding proteins (PBPs) for synthesis of 4→3 cross-links in the peptidoglycan of bacterial cell walls. Unusual 3→3 cross-links formed by L,D-transpeptidases were first detected in Escherichia coli more than four decades ago, however no phenotype has previously been associated with their synthesis. Here we show that production of the L,D-transpeptidase YcbB in combination with elevated synthesis of the (p)ppGpp alarmone by RelA lead to full bypass of the D,D-transpeptidase activity of PBPs and to broad-spectrum β-lactam resistance. Production of YcbB was therefore sufficient to switch the role of (p)ppGpp from antibiotic tolerance to high-level β-lactam resistance. This observation identifies a new mode of peptidoglycan polymerization in E. coli that relies on an unexpectedly small number of enzyme activities comprising the glycosyltransferase activity of class A PBP1b and the D,D-carboxypeptidase activity of DacA in addition to the L,D-transpeptidase activity of YcbB. DOI: http://dx.doi.org/10.7554/eLife.19469.001 PMID:27767957

  1. In Vitro and In Vivo Efficacy of β-Lactams against Replicating and Slowly Growing/Nonreplicating Mycobacterium tuberculosis

    PubMed Central

    Dinesh, Neela; Shandil, Radha; Ramachandran, Vasanthi; Sharma, Sreevalli; Bhattacharjee, Deepa; Ganguly, Samit; Reddy, Jitendar; Ahuja, Vijaykamal; Panduga, Vijender; Parab, Manish; Vishwas, K. G.; Kumar, Naveen; Balganesh, Meenakshi; Balasubramanian, V.

    2013-01-01

    Beta-lactams, in combination with beta-lactamase inhibitors, are reported to have activity against Mycobacterium tuberculosis bacteria growing in broth, as well as inside the human macrophage. We tested representative beta-lactams belonging to 3 different classes for activity against replicating M. tuberculosis in broth and nonreplicating M. tuberculosis under hypoxia, as well as against streptomycin-starved M. tuberculosis strain 18b (ss18b) in the presence or absence of clavulanate. Most of the combinations showed bactericidal activity against replicating M. tuberculosis, with up to 200-fold improvement in potency in the presence of clavulanate. None of the combinations, including those containing meropenem, imipenem, and faropenem, killed M. tuberculosis under hypoxia. However, faropenem- and meropenem-containing combinations killed strain ss18b moderately. We tested the bactericidal activities of meropenem-clavulanate and amoxicillin-clavulanate combinations in the acute and chronic aerosol infection models of tuberculosis in BALB/c mice. Based on pharmacokinetic/pharmacodynamic indexes reported for beta-lactams against other bacterial pathogens, a cumulative percentage of a 24-h period that the drug concentration exceeds the MIC under steady-state pharmacokinetic conditions (%TMIC) of 20 to 40% was achieved in mice using a suitable dosing regimen. Both combinations showed marginal reduction in lung CFU compared to the late controls in the acute model, whereas both were inactive in the chronic model. PMID:23507276

  2. In vitro and in vivo efficacy of β-lactams against replicating and slowly growing/nonreplicating Mycobacterium tuberculosis.

    PubMed

    Solapure, Suresh; Dinesh, Neela; Shandil, Radha; Ramachandran, Vasanthi; Sharma, Sreevalli; Bhattacharjee, Deepa; Ganguly, Samit; Reddy, Jitendar; Ahuja, Vijaykamal; Panduga, Vijender; Parab, Manish; Vishwas, K G; Kumar, Naveen; Balganesh, Meenakshi; Balasubramanian, V

    2013-06-01

    Beta-lactams, in combination with beta-lactamase inhibitors, are reported to have activity against Mycobacterium tuberculosis bacteria growing in broth, as well as inside the human macrophage. We tested representative beta-lactams belonging to 3 different classes for activity against replicating M. tuberculosis in broth and nonreplicating M. tuberculosis under hypoxia, as well as against streptomycin-starved M. tuberculosis strain 18b (ss18b) in the presence or absence of clavulanate. Most of the combinations showed bactericidal activity against replicating M. tuberculosis, with up to 200-fold improvement in potency in the presence of clavulanate. None of the combinations, including those containing meropenem, imipenem, and faropenem, killed M. tuberculosis under hypoxia. However, faropenem- and meropenem-containing combinations killed strain ss18b moderately. We tested the bactericidal activities of meropenem-clavulanate and amoxicillin-clavulanate combinations in the acute and chronic aerosol infection models of tuberculosis in BALB/c mice. Based on pharmacokinetic/pharmacodynamic indexes reported for beta-lactams against other bacterial pathogens, a cumulative percentage of a 24-h period that the drug concentration exceeds the MIC under steady-state pharmacokinetic conditions (%TMIC) of 20 to 40% was achieved in mice using a suitable dosing regimen. Both combinations showed marginal reduction in lung CFU compared to the late controls in the acute model, whereas both were inactive in the chronic model. PMID:23507276

  3. Validation study of the BetaStar plus lateral flow assay for detection of beta-lactam antibiotics in milk.

    PubMed

    Abouzied, Mohamed; Driksna, Dana; Walsh, Coilin; Sarzynski, Michael; Walsh, Aaron; Ankrapp, David; Klein, Frank; Rice, Jennifer; Mozola, Mark

    2012-01-01

    A validation study designed to meet the requirements of the AOAC Research Institute and the U.S. Food and Drug Administration, Center for Veterinary Medicine (FDA/CVM) was conducted for a receptor and antibody-based, immunochromatographic method (BetaStar Plus) for detection of beta-lactam antibiotic residues in raw, commingled bovine milk. The assay was found to detect amoxicillin, ampicillin, ceftiofur, cephapirin, cloxacillin, and penicillin G at levels below the FDA tolerance/safe levels, but above the maximum sensitivity thresholds established by the National Conference on Interstate Milk Shipments (NCIMS). Results of the part I (internal) and part II (independent laboratory) dose-response studies employing spiked samples were in close agreement. The test was able to detect all six drugs at the approximate 90/95% sensitivity levels when presented as incurred residues in milk collected from cows that had been treated with the specific drug. Selectivity of the assay was 100%, as no false-positive results were obtained in testing of 1031 control milk samples. Results of ruggedness experiments established the operating parameter tolerances for the BetaStar Plus assay. Results of cross-reactivity testing established that the assay detects certain other beta-lactam drugs (dicloxacillin and ticarcillin), but it does not cross-react with any of 30 drugs belonging to other classes. Abnormally high bacterial or somatic cell counts in raw milk produced no interference with the ability of the test to detect beta-lactams at tolerance/safe levels.

  4. Differential inactivation of alfalfa nodule glutamine synthetases by tabtoxinine-. beta. -lactam. [Pseudomonas syringae

    SciTech Connect

    Knight, T.J.; Unkefer, P.J.

    1987-04-01

    The presence of the pathogen Pseudomonas syringae pv. tabaci within the rhizosphere of nodulated alfalfa plants results in an increase in N/sub 2/-fixation potential and growth, but a 40-50% decrease in nodule glutamine synthetase (GS) activity, as compared to nodulated control plants. Tabtoxinine-..beta..-Lactam an exocellular toxin produced by Pseudomonas syringae pv tabaci irreversibly inhibits glutamine synthetase. Partial purification of nodule GS by DEAE-cellulose chromatography reveals two enzyme forms are present (GS/sub n1/ and GS/sub n2/). In vitro inactivation of the two glutamine synthetases associated with the nodule indicates a differential sensitivity to T-..beta..-L. The nodule specific GS/sub n1/ is much less sensitive to T-..beta..-L than the GS/sub n2/ enzyme, which was found to coelute with the root enzyme (GS/sub r/). However, both GS/sub n1/ and GS/sub n2/ are rapidly inactivated by methionine sulfoximine, another irreversible inhibitor of GS.

  5. Transcriptional induction of Streptomyces cacaoi beta-lactamase by a beta-lactam compound.

    PubMed

    Forsman, M; Lindgren, L; Häggström, B; Jaurin, B

    1989-10-01

    The soil bacterium Streptomyces cacaoi produces an extracellular beta-lactamase. The beta-lactamase expression could be induced by the beta-lactam compound 6-amino penicillinoic acid (6-APA). In liquid cultures, a 50-fold increase in beta-lactamase expression was observed within the first three hours after addition of 6-APA. Using the cloned beta-lactamase gene as a probe, it was shown that this increase was mediated at the level of transcriptional initiation. The start point of the induced beta-lactamase transcript was determined, and the nucleotide sequence of the promoter region was analysed. No noticeable homology was found to control regions of inducible beta-lactamase genes of other bacteria. A striking feature was the presence of six direct repeats (ten base pairs each) upstream of the promoter region. Thus, an example of an inducible regulatory gene system in this Gram-positive microorganism is presented. Also, the primary structure of the beta-lactamase was deduced, showing a high degree of homology with class A beta-lactamases. PMID:2559297

  6. A catalase-peroxidase for oxidation of β-lactams to their (R)-sulfoxides.

    PubMed

    Sangar, Shefali; Pal, Mohan; Moon, Lomary S; Jolly, Ravinder S

    2012-07-01

    In this communication we report for the first time a biocatalytic method for stereoselective oxidation of β-lactams, represented by penicillin-G, penicillin-V and cephalosporin-G to their (R)-sulfoxides. The method involves use of a bacterium, identified as Bacillus pumilis as biocatalyst. The enzyme responsible for oxidase activity has been purified and characterized as catalase-peroxidase (KatG). KatG of B. pumilis is a heme containing protein showing characteristic heme spectra with soret peak at 406 nm and visible peaks at 503 and 635 nm. The major properties that distinguish B. pumilis KatG from other bacterial KatGs are (i) it is a monomer and contains one heme per monomer, whereas KatGs of other bacteria are dimers or tetramers and have low heme content of about one per dimer or two per tetramer and (ii) its 12-residue, N-terminal sequence obtained by Edman degradation did not show significant similarity with any of known KatGs. PMID:21996477

  7. N-arylated-lactam-type iminosugars as new immunosuppressive agents: discovery, optimization, and biological evaluation.

    PubMed

    Wu, Xiaowei; Zhang, Fu-Yu; Zhu, Jingjing; Song, Chengcheng; Xiong, De-Cai; Zhou, Yifa; Cui, Yuxin; Ye, Xin-Shan

    2014-08-01

    We have previously described the discovery of N-alkylated iminosugars that showed immunosuppressive activity both in vitro and in vivo. Herein, we report the synthesis and biological evaluation of N-arylated lactam-type iminosugar derivatives. The synthesis started from simple monosaccharides and featured a Buchwald-Hartwig coupling reaction to construct the key N-aryl connection, thereby providing a highly diverse compound library. Structure-activity relationship studies, guided by a mouse-spleen-proliferation assay, led to the identification of 'hit' compound 12 f. Subsequently, the systematic modification of compound 12 f afforded compounds 21 h, 21 k, 21 n, 21 t, and 21 x with improved activities (IC50 =12-30 μM) and low Jurkat cytotoxicities (IC50 >100 μM). These new compounds also inhibited the secretion of IFN-γ and IL-4, which are hallmark cytokines of Th1 and Th2 cells, respectively. This work demonstrated that the N-arylated iminosugar structure represents a new scaffold with immunosuppressive activity.

  8. Chalcogen analogues of nicotine lactam studied by NMR, FTIR, DFT and X-ray methods

    NASA Astrophysics Data System (ADS)

    Jasiewicz, Beata; Malczewska-Jaskóła, Karolina; Kowalczyk, Iwona; Warżajtis, Beata; Rychlewska, Urszula

    2014-07-01

    The selenoanalogue of nicotine has been synthesized and characterized by spectroscopic and X-ray diffraction methods. The crystals of selenonicotine are isomorphic with the thionicotine homologue and consist of molecules engaged in columnar π⋯π stacking interactions between antiparallely arranged pyridine moieties. These interactions, absent in other crystals containing nicotine fragments, seem to be induced by the presence of a lactam group. The molecular structures in the vacuum of the oxo-, thio- and selenonicotine homologues have been calculated by the DFT method and compared with the available X-ray data. The delocalized structure of thionicotine is stabilized by intramolecular Csbnd H⋯S hydrogen bond, which becomes weaker in the partial zwitterionic resonance structure of selenonicotine in favor of multiple Csbnd H⋯Se intermolecular hydrogen-bonds. The calculated data allow a complete assignment of vibration modes in the solid state FTIR spectra. The 1H and 13C NMR chemical shifts were calculated by the GIAO method with B3LYP/6-311G(3df) level. A comparison between experimental and calculated theoretical results indicates that the density functional B3LYP method provided satisfactory results for predicting FTIR, 1H, 13C NMR spectra properties.

  9. Bioactivity-guided isolation of laevicarpin, an antitrypanosomal and anticryptococcal lactam from Piper laevicarpu (Piperaceae).

    PubMed

    da Silva A Maciel, Dayany; Freitas, Viviane P; Conserva, Geanne A Alves; Alexandre, Tatiana R; Purisco, Sonia U; Tempone, Andre G; Melhem, Márcia Souza C; Kato, Massuo J; Guimarães, Elsie F; Lago, João Henrique G

    2016-06-01

    Crude CH2Cl2 extract from leaves of Piper laevicarpu (Piperaceae) displayed antitrypanosomal activity against trypomastigote forms of Trypanosoma cruzi (Y strain) and antimicrobial potential against Cryptococcus gattii (strain-type WM 178). Bioactivity-guided fractionation of crude extract afforded one new natural bioactive lactam derivative, named laevicarpin. The structure of isolated compound, which displayed a very rare ring system, was elucidated based on NMR, IR and MS spectral analysis. Using MTT assay, the trypomastigotes of T. cruzi demonstrated susceptibility to laevicarpin displaying IC50 value of 14.7μg/mL (49.6μM), about 10-fold more potent than the standard drug benznidazole. The mammalian cytotoxicity of laevicarpin was verified against murine fibroblasts (NCTC cells) and demonstrated a CC50 value of 100.3μg/mL (337.7μM-SI=7). When tested against Cryptococcus gattii, laevicarpin showed an IC50 value of 2.3μg/mL (7.9μM) and a MIC value of 7.4μg/mL (25μM). Based in the obtained results, laevicarpin could be used as a scaffold for future drug design studies against the Chagas disease and anti-cryptococosis agents. PMID:27083380

  10. Poly-amido-saccharides: synthesis via anionic polymerization of a β-lactam sugar monomer.

    PubMed

    Dane, Eric L; Grinstaff, Mark W

    2012-10-01

    Enantiopure poly-amido-saccharides (PASs) with a defined molecular weight and narrow dispersity are synthesized using an anionic ring-opening polymerization of a β-lactam sugar monomer. The PASs have a previously unreported main chain structure that is composed of pyranose rings linked through the 1- and 2-positions by an amide with α-stereochemistry. The monomer is synthesized in one-step from benzyl-protected D-glucal and polymerized using mild reaction conditions to give degrees of polymerization ranging from 25 to >120 in high yield. Computational modeling reveals how the monomer's structure and steric bulk affect the thermodynamics and kinetics of polymerization. Protected and deprotected polymers and model compounds are characterized using a variety of methods (NMR, GPC, IR, DLS, etc.). On the basis of circular dichroism, the deprotected polymer possesses a regular secondary structure in aqueous solution, which agrees favorably with the prediction of a helical structure using molecular modeling. Furthermore, we provide evidence suggesting that the polymers bind the lectin concanavalin A at the same site as natural carbohydrates, showing the potential of these polymers to mimic natural polysaccharides. PASs offer the advantages associated with synthetic polymers, such as greater control over structure and derivitization. At the same time, they preserve many of the structural features of natural polysaccharides, such as a stereochemically regular, rigid pyranose backbone, that make natural carbohydrate polymers important materials both for their unique properties and useful applications.

  11. Physico-chemical characterization of a novel tricyclic beta-lactam antibiotic.

    PubMed

    Marini, A; Berbenni, V; Bruni, G; Sinistri, C; Maggioni, A; Orlandi, A; Villa, M

    2000-02-01

    GV118819X, a novel tricyclic beta-lactam antibiotic of GlaxoWellcome, is a racemic mixture of two diastereoisomers, A and B. Of the two diastereoisomers, only A is available as a pure compound. By analyzing mixtures of GV118819X and A, a partial phase diagram is constructed, which indicates the presence of a eutectic when the A fraction is approximately 39%. Moreover, the melting enthalpies of the eutectic mixture and of diastereoisomer B can be estimated. With the exception of the pure A form, all mixtures undergo important modifications in morphology and microstructure as a consequence of thermal treatments, which induce melting/amorphization of the eutectic, and crystallization of the A form. Analyses of the sieved fractions of GV118819X demonstrate that it consists of acicular crystals of different composition, with the larger crystals having a larger A fraction than the smaller ones. Grinding causes melting/amorphization of the eutectic and, following hours-long treatments, the formation of a substantial fraction of submicron particles with unusually low melting temperatures. PMID:10688752

  12. Aspects on evolution of fungal beta-lactam biosynthesis gene clusters and recruitment of trans-acting factors.

    PubMed

    Brakhage, Axel A; Thön, Marcel; Spröte, Petra; Scharf, Daniel H; Al-Abdallah, Qusai; Wolke, Sandra M; Hortschansky, Peter

    2009-01-01

    Penicillins and cephalosporins are beta-lactam antibiotics. The formation of hydrophobic penicillins has been reported in fungi only, notably Penicillium chrysogenum and Aspergillus (Emericella) nidulans, whereas the hydrophilic cephalosporins are produced by both fungi, e.g., Acremonium chrysogenum (cephalosporin C), and bacteria. The producing bacteria include Gram-negatives and Gram-positives, e.g., Streptomyces clavuligerus (cephamycin C) and Lysobacter lactamgenus (cephabacins), respectively. The evolutionary origin of beta-lactam biosynthesis genes has been the subject of discussion for many years, and two main hypotheses have been proposed: (i) horizontal gene transfer (HGT) from bacteria to fungi or (ii) vertical decent. There are strong arguments in favour of HGT, e.g., unlike most other fungal genes, beta-lactam biosynthesis genes are clustered and some of these genes lack introns. In contrast to S. clavuligerus, all regulators of fungal beta-lactam biosynthesis genes represent wide-domain regulators that are not part of the gene cluster. If bacterial regulators were co-transferred with the gene cluster from bacteria to fungi, most likely they would have been non-functional in eukaryotes and lost during evolution. Recently, the penicillin biosynthesis gene aatB was discovered, which is not part of the penicillin biosynthesis gene cluster and is even located on a different chromosome. The aatB gene is regulated by the same regulators AnCF and AnBH1 as the penicillin biosynthesis gene aatA (penDE). Data suggest that aatA and aatB are paralogues derived by duplication of a common ancestor gene. This data supports a model in which part of the beta-lactam biosynthesis gene cluster was transferred to some fungi, i.e., the acvA and ipnA gene without a regulatory gene. We propose that during the assembly of aatA and acvA-ipnA into a single gene cluster, recruitment of transcriptional regulators occurred along with acquisition of the duplicated aatA ancestor gene

  13. An investigation of resistance to β-lactam antimicrobials among staphylococci isolated from pigs with exudative epidermitis

    PubMed Central

    2013-01-01

    Background A high proportion of staphylococci isolated from pigs affected with exudative epidermitis were found to be resistant to β-lactam antimicrobials. The primary objective of this research was to investigate and characterize β-lactam resistance in Staphylococcus hyicus, Staphylococcus aureus and other staphylococci isolated from these pigs. Results The antimicrobial resistance patterns of 240 staphylococci isolates were determined by disk diffusion, of which 176 (73.3%) of the isolates were resistant to 3 β-lactams (penicillin G, ampicillin, and ceftiofur). The presence of mecA gene was identified in 63 staphylococci isolates from skin samples by PCR. The mecA gene was identified in 19 S. aureus, 31 S. hyicus, 9 Staphylococcus chromogenes, 2 Staphylococcus pseudintermedius isolates, and in 1 isolate each of Staphylococcus arlettae, and Staphylococcus cohnii subspecies urealyticus. From SCCmec typing results, the majority (45/63, 71.4%) were shown to be SCCmec type V. One isolate was SCCmec III. Fourteen isolates were detected as mec class A, mec class C or ccr type 5. The ccr complex and mec complex was not detected in 3 isolates of methicillin resistant S. hyicus (MRSH) based on multiplex PCR. Of the 30 isolates of MRSA identified from nasal samples of the pigs, 29 isolates were SCCmec type V and 1 isolate was SCCmec type II. Staphyloccoci isolates that were mecA negative but resistant to β-lactam antimicrobials were further examined by screening for mecC, however all were negative. Furthermore, the majority of mecA negative β-lactam resistant staphylococci isolates were susceptible to oxacillin and amoxicillin-clavulanic acid in a double disk diffusion test. Conclusions Methicillin resistance can be identified in a variety of staphylococcal species isolated from pigs. In this study there was a great deal of similarity in the SCCmec types between staphylococcal species, suggesting that resistance may be passed from one species of staphylococci to

  14. In Vivo Reversion to the Wild-Type β-Lactam Resistance Phenotype Mediated by a Plasmid Carrying ampR and qnrA1 in Enterobacter cloacae

    PubMed Central

    González-López, J. J.; Sabaté, M.; Lavilla, S.; Larrosa, M. N.; Bartolomé, R. M.; Prats, G.

    2006-01-01

    Resistance to β-lactams and quinolones in two isogenic Enterobacter cloacae isolates was studied. One was susceptible to cefoxitin and amoxicillin-clavulanate. The other one showed its natural β-lactam resistance pattern. Both isolates had a nonfunctional AmpR regulator. However, within the second one, the presence of a plasmid carrying ampR and qnrA1 allowed reversion to the wild-type β-lactam resistance phenotype and decreased susceptibility to fluoroquinolones. PMID:16940123

  15. Gene clusters for beta-lactam antibiotics and control of their expression: why have clusters evolved, and from where did they originate?

    PubMed

    Liras, Paloma; Martín, Juan F

    2006-03-01

    While beta-lactam compounds were discovered in filamentous fungi, actinomycetes and gram-negative bacteria are also known to produce different types of beta-lactams. All beta-lactam compounds contain a four-membered beta-lactam ring. The structure of their second ring allows these compounds to be classified into penicillins, cephalosporins, clavams, carbapenens or monobactams. Most beta-lactams inhibits bacterial cell wall biosynthesis but others behave as beta-lactamase inhibitors (e.g., clavulanic acid) and even as antifungal agents (e.g., some clavams). Due to the nature of the second ring in beta-lactam molecules, the precursors and biosynthetic pathways of clavams, carbapenems and monobactams differ from those of penicillins and cephalosporins. These last two groups, including cephamycins and cephabacins, are formed from three precursor amino acids that are linked into the alpha-aminoadipyl-L-cysteinyl-D-valine tripeptide. The first two steps of their biosynthetic pathways are common. The intermediates of these pathways, the characteristics of the enzymes involved, the lack of introns in the genes and bioinformatic analysis suggest that all of them should have evolved from an ancestral gene cluster of bacterial origin, which was surely transferred horizontally in the soil from producer to non-producer microorganisms. The receptor strains acquired fragments of the original bacterial cluster and occasionally inserted new genes into the clusters, which once modified, acquired new functions and gave rise to the final compounds that we know. When the order of genes in the Streptomyces genome is analyzed, the antibiotic gene clusters are highlighted as gene islands in the genome. Nonetheless, the assemblage of the ancestral beta-lactam gene cluster remains a matter of speculation. PMID:16636985

  16. Nitric Oxide from IFNγ-Primed Macrophages Modulates the Antimicrobial Activity of β-Lactams against the Intracellular Pathogens Burkholderia pseudomallei and Nontyphoidal Salmonella

    PubMed Central

    Jones-Carson, Jessica; Zweifel, Adrienne E.; Tapscott, Timothy; Austin, Chad; Brown, Joseph M.; Jones, Kenneth L.; Voskuil, Martin I.; Vázquez-Torres, Andrés

    2014-01-01

    Our investigations show that nonlethal concentrations of nitric oxide (NO) abrogate the antibiotic activity of β-lactam antibiotics against Burkholderia pseudomallei, Escherichia coli and nontyphoidal Salmonella enterica serovar Typhimurium. NO protects B. pseudomallei already exposed to β-lactams, suggesting that this diatomic radical tolerizes bacteria against the antimicrobial activity of this important class of antibiotics. The concentrations of NO that elicit antibiotic tolerance repress consumption of oxygen (O2), while stimulating hydrogen peroxide (H2O2) synthesis. Transposon insertions in genes encoding cytochrome c oxidase-related functions and molybdenum assimilation confer B. pseudomallei a selective advantage against the antimicrobial activity of the β-lactam antibiotic imipenem. Cumulatively, these data support a model by which NO induces antibiotic tolerance through the inhibition of the electron transport chain, rather than by potentiating antioxidant defenses as previously proposed. Accordingly, pharmacological inhibition of terminal oxidases and nitrate reductases tolerizes aerobic and anaerobic bacteria to β-lactams. The degree of NO-induced β-lactam antibiotic tolerance seems to be inversely proportional to the proton motive force (PMF), and thus the dissipation of ΔH+ and ΔΨ electrochemical gradients of the PMF prevents β-lactam-mediated killing. According to this model, NO generated by IFNγ-primed macrophages protects intracellular Salmonella against imipenem. On the other hand, sublethal concentrations of imipenem potentiate the killing of B. pseudomallei by NO generated enzymatically from IFNγ-primed macrophages. Our investigations indicate that NO modulates the antimicrobial activity of β-lactam antibiotics. PMID:25121731

  17. Poly-amido-saccharides: Synthesis via Anionic Polymerization of a β-Lactam Sugar Monomer

    PubMed Central

    Dane, Eric L.; Grinstaff, Mark W.

    2013-01-01

    Chiral poly-amido-saccharides (PASs) with a defined molecular weight and narrow polydispersity are synthesized using an anionic ring-opening polymerization of a β-lactam sugar monomer. The PASs have a previously unreported main chain structure that is composed of pyranose rings linked through the 1- and 2-positions by an amide bond with α-stereochemistry. The monomer is synthesized in one-step from benzyl-protected d-glucal and polymerized using mild reaction conditions to give degrees of polymerization ranging from 25 to >150 in high yield. Computational modeling reveals how the monomer’s structure and steric bulk affect the thermodynamics and kinetics of polymerization. Protected and deprotected polymers and model compounds are characterized using a variety of methods (NMR, GPC, IR, DLS, etc.). Reductive debenzylation provides the deprotected, hydrophilic polymers in high yield. Based on circular dichroism, the deprotected polymers possess a regular secondary structure in aqueous solution, which agrees favorably with the prediction of a helical structure using molecular modeling. Furthermore, we provide evidence suggesting that the polymers bind the lectin concanavalin A at the same site as natural carbohydrates, showing the potential of these polymers to mimic natural polysaccharides. PASs offer the advantages associated with synthetic polymers, such as greater control over structure and derivitization, and less batch-to-batch variation. At the same time, they preserve many of the structural features of natural polysaccharides, such as a stereochemically regular, rigid pyranose backbone, that make natural carbohydrate polymers important materials both for their unique properties and useful applications. PMID:22937875

  18. Transcriptional control of expression of fungal beta-lactam biosynthesis genes.

    PubMed

    Litzka, O; Then Bergh, K; Van den Brulle, J; Steidl, S; Brakhage, A A

    1999-01-01

    The most commonly used beta-lactam antibiotics for the therapy of infectious diseases are penicillin and cephalosporin. Penicillin is produced as end product by some fungi most notably by Aspergillus (Emericella) nidulans and Penicillium chrysogenum. Cephalosporins are synthesised by several bacteria and fungi, e.g. by the fungus Acremonium chrysogenum (syn. Cephalosporium acremonium). The biosynthetic pathways leading to both secondary metabolites start from the same three amino acid precursors and have the first two enzymatic reactions in common. The penicillin biosynthesis is catalysed by three enzymes encoded by acvA (pcbAB), ipnA (pcbC) and aatA (penDE). The genes are organised into a cluster. In A. chrysogenum, in addition to acvA and ipnA, which are also clustered, a second cluster contains the genes for enzymes catalysing the reactions of the later steps of the cephalosporin pathway (cefEF, cefG). Transcription of biosynthesis genes is subject to sophisticated control by nutritional factors (e.g. glucose, nitrogen), amino acids such as lysine and methionine, and ambient pH. Some regulators have been identified such as the A. nidulans pH regulatory protein PACC and the transcriptional complex PENR1. PENR1 is a HAP-like transcriptional complex similar or identical to AnCF. Additional positive regulatory factors seem to be represented by recessive trans-acting mutations of A. nidulans (prgA1, prgB1, npeE1) and P. chrysogenum (carried by mutants Npe2 and Npe3). The GATA-binding factor NRE appears to be involved in the regulation of the penicillin biosynthesis genes by the nitrogen source in P. chrysogenum. Formal genetic evidence suggests the existence of transcriptional repressors as well.

  19. Saliva, supragingival biofilm and root canals can harbor gene associated with resistance to lactamic agents.

    PubMed

    Moraes, Ludmila Coutinho; Fatturi-Parolo, Clarissa Cavalcanti; Ferreira, Maria Beatriz Cardoso; Só, Marcus Vinicius Reis; Montagner, Francisco

    2015-01-01

    This study aimed to determine the presence of Prevotella strains and genes associated with resistance to lactamics in different oral niches from patients with/without primary endodontic infections. Saliva (S) and supragingival biofilm (SB) were collected from three patient groups: Group I - no endodontic infection (n = 15); Group II - acute endodontic infection (n = 12); and Group III - chronic endodontic infection (n = 15). Root canal (RC) samples were collected from Groups II and III. The presence of P. intermedia, P nigrescens, P. tannerae and cfxA/cfxA2 gene was assessed by PCR. The cfxA/cfxA2 gene was not detected in all environments within the same patient. The cfxA/cfxA2 gene was present in 23.81% of S samples, 28.57% of SB samples, and 7.41% of RC samples. Prevotella species were detected in 53.97%, 47.62% and 34.56% of the S, SB, and RC samples, respectively. P. intermedia had a high frequency in saliva samples from Group 3. Saliva samples from Group 1 had higher detection rates of P. nigrescens than did Groups 2 and 3. Patients without endodontic disease had high frequencies of P. nigrescens in the SB samples. The presence or absence of spontaneous symptoms was not related to the detection rates for resistance genes in the RC samples. Saliva, supragingival biofilm and root canals can harbor resistant bacteria. The presence of symptomatology did not increase the presence of the cfxA/cfxA2 gene in the supragingival biofilm and inside root canals. PMID:25789508

  20. Diversity of mechanisms conferring resistance to β-lactams among OXA-23-producing Acinetobacter baumannii clones.

    PubMed

    Cardoso, Juliana Provasi; Cayô, Rodrigo; Girardello, Raquel; Gales, Ana Cristina

    2016-05-01

    A total of 31 unrelated OXA-23-producing Acinetobacter baumannii strains isolated from 14 hospitals located in distinct Brazilian regions were evaluated in this study. These isolates were grouped into 12 different sequence types (STs), of which 7 had unique allelic sequences (ST188, ST189, ST190, ST191, ST192, ST228, and ST299). Most isolates belonged to the clonal complex CC79 followed by CC15 and CC1. Only polymyxin B and minocycline showed good activity against the OXA-23-producing A. baumannii clones. The ISAba1 upstream blaOXA-23, blaOXA-51-like, or ampC was found in 100%, 54.8%, and 77.4% of the isolates, respectively. High resistance rates to ceftazidime and cefotaxime were observed among those isolates possessing ISAba1 upstream ampC, in contrast to those isolates that did not carry this configuration. Moreover, a ≥2 Log2 decrease in the MICs of meropenem and ceftazidime was observed in the presence of phenyl-arginine-β-naphthylamide for 80.6% and 54.8% of isolates, respectively. Overexpression of the adeB was observed in 61.3% of isolates, particularly among those isolates belonging to the ST1 (CC1). It was also verified that ompW was down-regulated in all isolates belonging to the ST15 (CC15). On the other hand, carO and omp33-36 genes were overexpressed in 48.4% and 58.1% of the isolates, respectively. In this study, we show that overexpression of AdeABC system could significantly contribute for resistance to meropenem and ceftazidime among OXA-23-producing A. baumannii clones in Brazil, demonstrating the complexity involved in the β-lactam resistance in such isolates. PMID:26971181

  1. Longitudinal surveillance of outpatient β-lactam antimicrobial use in Canada, 1995 to 2010

    PubMed Central

    Glass-Kaastra, Shiona K; Finley, Rita; Hutchinson, Jim; Patrick, David M; Weiss, Karl; Conly, John

    2014-01-01

    INTRODUCTION: β-lactam antimicrobials are the most commonly prescribed group of antimicrobials in Canada, and are categorized by the WHO as critically and highly important antimicrobials for human medicine. Because antimicrobial use is commonly associated with the development of antimicrobial resistance, monitoring the volume and patterns of use of these agents is highly important. OBJECTIVE: To assess the use of penicillin and cephalosporin antimicrobials within Canadian provinces over the 1995 to 2010 time frame according to two metrics: prescriptions per 1000 inhabitant-days and the average defined daily doses dispensed per prescription. METHODS: Antimicrobial prescribing data were acquired from the Canadian Integrated Program for Antimicrobial Resistance Surveillance and the Canadian Committee for Antimicrobial Resistance, and population data were obtained from Statistics Canada. The two measures developed were used to produce linear mixed models to assess differences among provinces and over time for the broad-spectrum penicillin and cephalosporin groups, while accounting for repeated measurements at the provincial level. RESULTS: Significant differences among provinces were found, as well as significant changes in use over time. A >28% reduction in broad-spectrum penicillin prescribing occurred in each province from 1995 to 2010, and a >18% reduction in cephalosporin prescribing occurred in all provinces from 1995 to 2010, with the exception of Manitoba, where cephalosporin prescribing increased by 18%. DISCUSSION: Significant reductions in the use of these important drugs were observed across Canada from 1995 to 2010. Newfoundland and Labrador and Quebec emerged as divergent from the remaining provinces, with high and low use, respectively. PMID:24855480

  2. Paper analytical devices for fast field screening of beta lactam antibiotics and anti-tuberculosis pharmaceuticals

    PubMed Central

    Weaver, Abigail A.; Reiser, Hannah; Barstis, Toni; Benvenuti, Michael; Ghosh, Debarati; Hunckler, Michael; Joy, Brittney; Koenig, Leah; Raddell, Kellie; Lieberman, Marya

    2013-01-01

    Reports of low quality pharmaceuticals have been on the rise in the last decade with the greatest prevalence of substandard medicines in developing countries, where lapses in manufacturing quality control or breaches in the supply chain allow substandard medicines to reach the marketplace. Here, we describe inexpensive test cards for fast field screening of pharmaceutical dosage forms containing beta lactam antibiotics or combinations of the four first-line antituberculosis (TB) drugs. The devices detect the active pharmaceutical ingredients (APIs) ampicillin, amoxicillin, rifampicin, isoniazid, ethambutol, and pyrazinamide, and also screen for substitute pharmaceuticals such as acetaminophen and chloroquine that may be found in counterfeit pharmaceuticals. The tests can detect binders and fillers like chalk, talc, and starch not revealed by traditional chromatographic methods. These paper devices contain twelve lanes, separated by hydrophobic barriers, with different reagents deposited in the lanes. The user rubs some of the solid pharmaceutical across the lanes and dips the edge of the paper into water. As water climbs up the lanes by capillary action, it triggers a library of different chemical tests and a timer to indicate when the tests are completed. The reactions in each lane generate colors to form a “color bar code” which can be analyzed visually by comparison to standard outcomes. While quantification of the APIs is poor compared to conventional analytical methods, the sensitivity and selectivity for the analytes is high enough to pick out suspicious formulations containing no API or a substitute API, as well as formulations containing APIs that have been “cut” with inactive ingredients. PMID:23725012

  3. N-Thiolated β-Lactams: Studies on the Mode of Action and Identification of a Primary Cellular Target in S. aureus

    PubMed Central

    Revell, Kevin D.; Heldreth, Bart; Long, Timothy E.; Jang, Seyoung; Turos, Edward

    2007-01-01

    This study focuses on the mechanism of action of N-alkylthio β-lactams, a new family of antibacterial compounds that show promising activity against Staphylococcus and Bacillus microbes. Previous investigations have determined that these compounds are highly selective towards which bacteria they target, and possess completely unprecedented structure-activity profiles for a β-lactam antibiotic. Unlike penicillin, which inhibits cell wall crosslinking proteins and affords a broad spectrum of bacteriocidal activity, these N-thiolated lactams are bacteriostatic in their behavior and act through a different mechanistic mode. Our current findings indicate that the compounds react rapidly within the bacterial cell with co-enzyme A (CoA) through in vivo transfer of the N-thio group to produce an alkyl-CoA mixed disulfide species, which then interferes with fatty acid biosynthesis. Our studies on coenzyme A disulfide reductase show that the CoA thiol redox buffer is not perturbed by these compounds; however, the lactams appear to act as prodrugs. The experimental evidence that these β-lactams inhibit fatty acid biosynthesis in bacteria, and the elucidation of coenzyme A as a primary cellular target, offers opportunities for the discovery of other small organic compounds that can be developed as therapeutics for MRSA and anthrax infections. PMID:17258460

  4. In vitro activities of 22 beta-lactam antibiotics against penicillin-resistant and penicillin-susceptible viridans group streptococci isolated from blood.

    PubMed Central

    Alcaide, F; Liñares, J; Pallares, R; Carratala, J; Benitez, M A; Gudiol, F; Martin, R

    1995-01-01

    A total of 410 strains of viridans group streptococci isolated consecutively from blood were tested by the microdilution method for in vitro susceptibility to 22 beta-lactam antibiotics. One hundred thirty-eight strains (33.6%) were resistant to penicillin with a MIC range of 0.25 to 8 micrograms/ml. MICs of all beta-lactam agents tested were higher for penicillin-resistant strains than for susceptible strains. These antibiotics were classified into three groups according to their in vitro activities (MICs at which 50 and 90% of the isolates are inhibited). Beta-Lactams of the first group (these included imipenem, cefpirome, FK-037, cefditoren, cefotaxime, ceftriaxone, and cefepime) showed activities higher than or similar to that of penicillin against penicillin-resistant viridans group streptococci. However, 80% of highly penicillin-resistant Streptococcus mitis organisms required cefotaxime and ceftriaxone MICs of > or = 2 micrograms/ml (range, 2 to 16 micrograms/ml). Beta-Lactams of the second group (cefpodoxime, ampicillin, amoxicillin-clavulanate, piperacillin, and cefuroxime) showed lower activities than penicillin. Finally, antibiotics of the third group (cephalothin, oxacillin, ceftazidime, cefixime, cefaclor, cefetamet, cefadroxil, cephalexin, and ceftibuten) showed poor in vitro activities. Therefore, some of the beta-lactam agents included in the first group could be an acceptable alternative in the treatment of serious infections due to strains highly resistant to penicillin, although clinical experience is needed. PMID:8619576

  5. Comprehensive allergy work-up is mandatory in cystic fibrosis patients who report a history suggestive of drug allergy to beta-lactam antibiotics

    PubMed Central

    2012-01-01

    Background In the general population, reports on suspected ß-lactam hypersensitivity are common. After a drug allergy work-up at best 20% of the selected patients are positive. However, these considerations have not been explored in cystic fibrosis patients for whom antibiotics are even more crucial. Methods The study, part of the Drug Allergy and Hypersensitivity (DAHD) cohort, was performed in the regional cystic fibrosis center of Montpellier, France. After identifying patients with a clinical history suggestive of drug allergy to ß-lactams, a complete drug allergy work-up, was carried out according to the EAACI recommendations. Results Among the 171 patients involved, 23 reported clinical manifestations potentially compatible with a drug allergy to ß-lactams. After performing the complete drug-allergy work-up, 7 were considered as drug hypersensitive (3 had positive skin tests, 1 a positive provocation test, 3 declined the tests). Excluding the latter 3 patients with incomplete drug allergy work-up, the rate of proven drug allergy was 2.3%. Conclusions Drug allergy to ß-lactams in cystic fibrosis patients is of importance. A full drug allergy work-up is mandatory in case of suspicion, because ß-lactam responsibility is often ruled out. PMID:22697261

  6. Evaluation of the Osiris expert system for identification of beta-lactam phenotypes in isolates of Pseudomonas aeruginosa.

    PubMed

    Bert, Frédéric; Ould-Hocine, Zahia; Juvin, Manette; Dubois, Véronique; Loncle-Provot, Véronique; Lefranc, Valérie; Quentin, Claudine; Lambert, Nicole; Arlet, Guillaume

    2003-08-01

    Osiris is a video zone size reader for disk diffusion tests featuring a built-in extended expert system (EES). The efficacy of the EES for the identification of the beta-lactam susceptibility phenotypes of Pseudomonas aeruginosa isolates was evaluated. Thirteen beta-lactams were tested in four laboratories by the disk diffusion test with 53 strains with well-characterized resistance mechanisms, including the production of 12 extended-spectrum beta-lactamases (ESBLs). The plates were read with the Osiris system and the results were interpreted with the ESS, and then the phenotype identified by the EES was compared to the resistance mechanism. The strains were also screened for the presence of ESBL production by a double-disk synergy test by placing the strains between an extended-spectrum cephalosporin-containing disk and a clavulanic acid-containing disk at distances of 30, 20, 15, and 10 mm from each other. Overall, the EES accurately identified the phenotypes of 88.2% of the strains and indicated an association with several mechanisms for 3.8% of the strains. No phenotype was identified in four strains with low levels of penicillinase production. Misidentifications were observed for two penicillinase-producing strains: one strain with partially derepressed cephalosporinase production and one strain overexpressing the MexA-MexB-OprM efflux system. The production of only four ESBLs was detected by the standard synergy test with a 30-mm distance between the disks. The production of five further ESBLs was identified by reducing the distance to 20 mm, and the production of the last three ESBLs was detected only at a distance of 15 or 10 mm. Our results indicate that the Osiris EES is an effective tool for the identification of P. aeruginosa beta-lactam phenotypes. A specific double-disk synergy test with reduced disk distances is necessary for the detection of ESBL production by this organism.

  7. Evaluation and updating of the Osiris expert system for identification of Escherichia coli beta-lactam resistance phenotypes.

    PubMed

    Bert, Frédéric; Juvin, Manette; Ould-Hocine, Zahia; Clarebout, Gervais; Keller, Emmanuelle; Lambert, Nicole; Arlet, Guillaume

    2005-04-01

    Osiris is a video zone size reader for disk diffusion tests that includes a built-in extended expert system (EES). We evaluated the efficacy of the Osiris EES for the identification of beta-lactam susceptibility phenotypes of Escherichia coli isolates. Fifteen beta-lactam agents and three beta-lactam-beta-lactamase inhibitor combinations were tested by the disk diffusion method against 50 E. coli strains with well-characterized resistance mechanisms. The strains were screened for the production of extended-spectrum beta-lactamase (ESBL) by the double-disk synergy test using a disk of amoxicillin-clavulanic acid with disks of the extended-spectrum cephalosporins and aztreonam. Overall, the EES accurately identified the phenotype for 78% of the strains, indicated an inexact phenotype for 17% of the strains, and could not find a matching phenotype for the remaining 5% of the strains. The percentage of correct identification for each resistance mechanism was 100% for inhibitor-resistant TEM and for TEM plus cephalosporinase, 88.9% for TEM and for ESBL, 70.8% for cephalosporinase overproduction, and 25% for oxacillinase. The main cause of discrepancy was the misidentification of oxacillinase as inhibitor-resistant TEM. The conventional double-disk synergy test failed to detect ESBL production in two strains (one producing VEB-1 and one producing CTX-M-14), but synergy between cefepime and amoxicillin-clavulanic acid was visible after the distance between the disks was reduced to 20 mm. After the interpretative guidelines of the EES were updated according to our results, the percentage of correct phenotype identification increased from 78 to 96%.

  8. Validation study of a receptor-based lateral flow assay for detection of beta-lactam antibiotics in milk.

    PubMed

    Abouzied, Mohamed; Sarzynski, Michael; Walsh, Aaron; Wood, Heather; Mozola, Mark

    2009-01-01

    A validation study designed to meet the requirements of the AOAC Research Institute and the U.S. Food and Drug Administration (FDA), Center for Veterinary Medicine, was conducted for a receptor-based, immunochromatographic method (BetaStar US) for detection of beta-lactam antibiotic residues in raw, commingled bovine milk. The assay was found to detect amoxicillin, ampicillin, cephapirin, cloxacillin, and penicillin G at levels below the FDA tolerance/safe levels but above the maximum sensitivity thresholds established by the National Conference on Interstate Milk Shipments. Results of the Part I (internal) and Part II (independent laboratory) dose-response studies using spiked samples were in very close agreement for all five drugs tested, with differences between the Part I and Part II 90/95% sensitivity values ranging from 0 to 1 ppb. The test was able to detect all five drugs at the approximate 90/95% sensitivity levels when present as incurred residues in milk collected from cows that had been treated with the specific drug. A sixth drug, ceftiofur, was found to be undetectable at levels of < or = 500 ppb (as total ceftiofur metabolites from incurred residues in milk samples). The selectivity of the assay was 100%, because no false-positive results were obtained in tests of >1000 control milk samples. The assay was found to be applicable to the testing of frozen raw milk samples. Results of ruggedness experiments established the operating parameter tolerances for the BetaStar US assay. Results of cross-reactivity testing established that the assay detects certain other beta-lactam drugs (dicloxacillin and ticarcillin), but it does not cross-react with any of 30 drugs belonging to other classes. Abnormally high bacterial or somatic cell counts in raw milk produced no interference with the ability of the test to detect beta-lactams at tolerance/safe levels.

  9. Transition Metal-Free Selective Double sp(3) C-H Oxidation of Cyclic Amines to 3-Alkoxyamine Lactams.

    PubMed

    Osorio-Nieto, Urbano; Chamorro-Arenas, Delfino; Quintero, Leticia; Höpfl, Herbert; Sartillo-Piscil, Fernando

    2016-09-16

    The first chemical method for selective dual sp(3) C-H functionalization at the alpha-and beta positions of cyclic amines to their corresponding 3-alkoxyamine lactams is reported. Unlike traditional Cα-H oxidation of amines to amides mediated by transition metals, the present protocol, which involves the use of NaClO2/TEMPO/NaClO in either aqueous or organic solvent, not only allows the Cα-H oxidation but also the subsequent functionalization of the unreactive β-methylene group in an unprecedented tandem fashion and using environmentally friendly reactants.

  10. Release of lipoteichoic acid from Staphylococcus aureus by treatment with cefmetazole and other beta-lactam antibiotics.

    PubMed

    Utsui, Y; Ohya, S; Takenouchi, Y; Tajima, M; Sugawara, S; Deguchi, K; Suginaka, H

    1983-10-01

    The effect of cefmetazole on the growth together with the release of cellular lipoteichoic acid from cefazolin-resistant strains of Staphylococcus aureus was compared with that of cefazolin, cefotiam, cefoxitin and cefuroxime. Bacteriolytic actions were measured by turbidity and bactericidal actions were followed by viable cell count. Release of cellular lipoteichoic acid was measured by the radioactivity in the supernatant of the cultures. Cefmetazole exerted more potent effects on the bacterial growth and induced more marked release of cellular lipoteichoic acid from resistant strains as compared with other beta-lactams.

  11. Transition Metal-Free Selective Double sp(3) C-H Oxidation of Cyclic Amines to 3-Alkoxyamine Lactams.

    PubMed

    Osorio-Nieto, Urbano; Chamorro-Arenas, Delfino; Quintero, Leticia; Höpfl, Herbert; Sartillo-Piscil, Fernando

    2016-09-16

    The first chemical method for selective dual sp(3) C-H functionalization at the alpha-and beta positions of cyclic amines to their corresponding 3-alkoxyamine lactams is reported. Unlike traditional Cα-H oxidation of amines to amides mediated by transition metals, the present protocol, which involves the use of NaClO2/TEMPO/NaClO in either aqueous or organic solvent, not only allows the Cα-H oxidation but also the subsequent functionalization of the unreactive β-methylene group in an unprecedented tandem fashion and using environmentally friendly reactants. PMID:27564379

  12. Regulation of the expression of the β-lactam antibiotic-resistance determinants in methicillin-resistant Staphylococcus aureus (MRSA).

    PubMed

    Blázquez, Blas; Llarrull, Leticia I; Luque-Ortega, Juan R; Alfonso, Carlos; Boggess, Bill; Mobashery, Shahriar

    2014-03-18

    β-Lactam antibiotics have faced obsolescence with the emergence of methicillin-resistant Staphylococcus aureus (MRSA). A complex set of events ensues upon exposure of MRSA to these antibiotics, which culminates in proteolysis of BlaI or MecI, two gene repressors, and results in the induction of resistance. We report studies on the mechanism of binding of these gene repressors to the operator regions by fluorescence anisotropy. Within the range of in vivo concentrations for BlaI and MecI, these proteins interact with their regulatory elements in a reversible manner, as both a monomer and a dimer. PMID:24564530

  13. Colletotrilactam A-D, novel lactams from Colletotrichum gloeosporioides GT-7, a fungal endophyte of Uncaria rhynchophylla.

    PubMed

    Wei, Bo; Yang, Zhong-Duo; Chen, Xiao-Wei; Zhou, Shuang-Yan; Yu, Hai-Tao; Sun, Jing-Yun; Yao, Xiao-Jun; Wang, Yong-Gang; Xue, Hong-Yan

    2016-09-01

    Four novel lactams, colletotrilactam A-D (1-4), along with six known compounds (5-10) were isolated from the culture broth of Colletotrichum gloeosporioides GT-7, a fungal endophyte of Uncaria rhynchophylla. The structures of these compounds were elucidated by comprehensive NMR spectroscopy. Isolates were tested for monoamine oxidase (MAO) inhibitory activity and compound 9 showed potent MAO inhibitory activity with IC50 value of 8.93±0.34μg/mL, when the IC50 value of iproniazid as a standard was 1.80±0.5μg/mL. PMID:27520493

  14. Highly efficient asymmetric hydrogenation of cyano-substituted acrylate esters for synthesis of chiral γ-lactams and amino acids.

    PubMed

    Kong, Duanyang; Li, Meina; Wang, Rui; Zi, Guofu; Hou, Guohua

    2016-01-28

    A highly efficient and enantioselective synthesis of γ-lactams and γ-amino acids by Rh-catalyzed asymmetric hydrogenation has been developed. Using the Rh-(S,S)-f-spiroPhos complex, under mild conditions a wide range of 3-cyano acrylate esters including both E and Z-isomers and β-cyano-α-aryl-α,β-unsaturated ketones were first hydrogenated with excellent enantioselectivities (up to 98% ee) and high turnover numbers (TON up to 10,000). PMID:26661067

  15. Choice of an oral beta-lactam antibiotic for infections due to penicillin-resistant Streptococcus pneumoniae.

    PubMed

    Goldstein, F W

    1997-01-01

    The activity, pharmacokinetic and pharmacodynamic parameters of 5 oral beta-lactams have been compared against 400 penicillin-resistant Streptococcus pneumoniae (PRSP) isolated in 9 European countries. All the data have been calculated for the highest recommended oral dosage in France. Amoxicillin was by far the most effective antibiotic when considering its intrinsic activity (96% of the PRSP inhibited at 2 mg/l), the inhibitory quotient (100% of the PRSP > 1 vs 69.2% for cefuroxime) or a time above MIC > 40% which best correlates with clinical results (96% for amoxicillin vs < 25% for the cephalosporins).

  16. •OH and e-aq are yet good candidates for demolishing the β-lactam system of a penicillin eliminating the antimicrobial activity

    NASA Astrophysics Data System (ADS)

    Szabó, László; Tóth, Tünde; Rácz, Gergely; Takács, Erzsébet; Wojnárovits, László

    2016-07-01

    Tracking the pharmacophore of a drug subjected to advanced oxidation is essential for evaluating the efficiency of the process in terms of wastewater treatment. From this standpoint, the •OH and eaq- induced deactivation mechanism of amoxicillin, a penicillin derivative was investigated in dilute aqueous solution using pulse- and gamma-radiolysis techniques. Based on IR measurements, •OH and eaq- destroys the β-lactam system of amoxicillin with ~55% and ~84% efficiency, respectively. In aerated solution the elimination of the pharmacophore was slightly impaired since the reaction pathway of the ring-opening was disturbed owing to the reactivity of O2 and O2• - toward the intermediates of sulfur oxidation. The high potency of eaq- for β-lactam deactivation is attributed to the enhanced electron deficiency of the carbonyl carbon inside the lactam ring.

  17. •OH and e-aq are yet good candidates for demolishing the β-lactam system of a penicillin eliminating the antimicrobial activity

    NASA Astrophysics Data System (ADS)

    Szabó, László; Tóth, Tünde; Rácz, Gergely; Takács, Erzsébet; Wojnárovits, László

    2016-07-01

    Tracking the pharmacophore of a drug subjected to advanced oxidation is essential for evaluating the efficiency of the process in terms of wastewater treatment. From this standpoint, the •OH and eaq- induced deactivation mechanism of amoxicillin, a penicillin derivative was investigated in dilute aqueous solution using pulse- and gamma-radiolysis techniques. Based on IR measurements, •OH and eaq- destroys the β-lactam system of amoxicillin with ~55% and ~84% efficiency, respectively. In aerated solution the elimination of the pharmacophore was slightly impaired since the reaction pathway of the ring-opening was disturbed owing to the reactivity of O2 and O2• - toward the intermediates of sulfur oxidation. The high potency of eaq- for β-lactam deactivation is attributed to the enhanced electron deficiency of the carbonyl carbon inside the lactam ring.

  18. Reaction of soluble penicillin-binding protein 2a of methicillin-resistant Staphylococcus aureus with beta-lactams and acyclic substrates: kinetics in homogeneous solution.

    PubMed Central

    Graves-Woodward, K; Pratt, R F

    1998-01-01

    The kinetics of reaction of solubilized penicillin-binding protein 2a (sPBP2a) of methicillin-resistant Staphylococcus aureus with a variety of beta-lactams and acyclic species was studied in homogeneous aqueous solution at 37 degreesC in 25 mM Hepes buffer, pH7.0, containing 1 M NaCl. Under these conditions, but not at lower salt concentrations, protein precipitation did not occur either during or after the reaction. The reactions of beta-lactams in general could be monitored by competition with a chromophoric beta-lactam, nitrocefin, or directly in certain cases by protein fluorescence. Rate constants for reaction of a wide variety of beta-lactams are reported. The interactions are characterized by a slow second-order acylation reaction followed by a slower deacylation. For example, the rate constants for benzylpenicillin were 12 M-1.s-1 and 3x10(-5) s-1 respectively. The acylation is slow in comparison with those of normal non-resistant high-molecular-mass penicillin-binding proteins. sPBP2a also seemed to catalyse the slow hydrolysis of a variety of acyclic depsipeptides but not that of a d-Ala-d-Ala peptide. The reactions with certain depsipeptides also led to protein precipitation. These reactions were, however, not affected by prior blockage of the beta-lactam-binding site by benzylpenicillin and thus might take place elsewhere on the enzyme. Two classes of potential transition- state analogue inhibitors, phosphonate monoesters and boronates, seemed to have little effect on the rate of reaction of sPBP2a with nitrocefin and therefore seem to have little affinity for the beta-lactam-binding/D,D-peptidase site. PMID:9620879

  19. Combinations of β-Lactam Antibiotics Currently in Clinical Trials Are Efficacious in a DHP-I-Deficient Mouse Model of Tuberculosis Infection

    PubMed Central

    Rullas, Joaquín; Dhar, Neeraj; McKinney, John D.; García-Pérez, Adolfo; Lelievre, Joël; Diacon, Andreas H.; Hugonnet, Jean-Emmanuel; Arthur, Michel; Angulo-Barturen, Iñigo; Barros-Aguirre, David

    2015-01-01

    We report here a dehydropeptidase-deficient murine model of tuberculosis (TB) infection that is able to partially uncover the efficacy of marketed broad-spectrum β-lactam antibiotics alone and in combination. Reductions of up to 2 log CFU in the lungs of TB-infected mice after 8 days of treatment compared to untreated controls were obtained at blood drug concentrations and time above the MIC (T>MIC) below clinically achievable levels in humans. These findings provide evidence supporting the potential of β-lactams as safe and mycobactericidal components of new combination regimens against TB with or without resistance to currently used drugs. PMID:25987618

  20. Zinc Finger Nuclease: A New Approach to Overcome Beta-Lactam Antibiotic Resistance

    PubMed Central

    Shahbazi Dastjerdeh, Mansoureh; Kouhpayeh, Shirin; Sabzehei, Faezeh; Khanahmad, Hossein; Salehi, Mansour; Mohammadi, Zahra; Shariati, Laleh; Hejazi, Zahra; Rabiei, Parisa; Manian, Mostafa

    2016-01-01

    Background: The evolution of antibiotic-resistant bacteria (ARB) and antibiotic-resistance genes (ARGs) has been accelerated recently by the indiscriminate application of antibiotics. Antibiotic resistance has challenged the success of medical interventions and therefore is considered a hazardous threat to human health. Objectives: The present study aimed to describe the use of zinc finger nuclease (ZFN) technology to target and disrupt a plasmid-encoded β-lactamase, which prevents horizontal gene transfer-mediated evolution of ARBs. Materials and Methods: An engineered ZFN was designed to target a specific sequence in the ampicillin resistance gene (ampR) of the pTZ57R plasmid. The Escherichia coli bacteria already contained the pZFN kanamycin-resistant (kanaR) plasmid as the case or the pP15A, kanaR empty vector as the control, were transformed with the pTZ57R; the ability of the designed ZFN to disrupt the β-lactamase gene was evaluated with the subsequent disturbed ability of the bacteria to grow on ampicillin (amp) and ampicillin-kanamycin (amp-kana)-containing media. The effect of mild hypothermia on the ZFN gene targeting efficiency was also evaluated. Results: The growth of bacteria in the case group on the amp and amp-kana-containing media was significantly lower compared with the control group at 37°C (P < 0.001). Despite being more efficient in hypothermic conditions at 30°C (P < 0.001), there were no significant associations between the incubation temperature and the ZFN gene targeting efficiency. Conclusions: Our findings revealed that the ZFN technology could be employed to overcome ampicillin resistance by the targeted disruption of the ampicillin resistance gene, which leads to inactivation of β-lactam synthesis. Therefore, ZFN technology could be engaged to decrease the antibiotic resistance issue with the construction of a ZFN archive against different ARGs. To tackle the resistance issue at the environmental level, recombinant phages

  1. Synthesis and fungistatic activity of bicyclic lactones and lactams against Botrytis cinerea, Penicillium citrinum, and Aspergillus glaucus.

    PubMed

    Walczak, Paulina; Pannek, Jakub; Boratyński, Filip; Janik-Polanowicz, Agata; Olejniczak, Teresa

    2014-08-27

    Six analogues of natural trans-4-butyl-cis-3-oxabicyclo[4.3.0]nonan-2-one (3) and three derivatives, 11, 12, and 13, of Vince lactam (10) were synthesized and tested as fungistatic agents against Botrytis cinerea AM235, Penicillium citrinum AM354, and six strains of Aspergillus. Moreover, bioresolution carried out by means of whole cell microorganisms and commercially available enzymes afforded opposite enantiomerically enriched (-) and (+) isomers of Vince lactam (10), respectively. The effect of compound structures and stereogenic centers on biological activity has been discussed. The highest fungistatic activity was observed for four lactones: 3, 4, 7, and 8 (IC50 = 104.6-115.2 μg/mL) toward B. cinerea AM235. cis-5,6-Epoxy-2-aza[2.2.1]heptan-3-one (13) indicated significant fungistatic activity (IC50 = 107.1 μg/mL) against Aspergillus glaucus AM211. trans-4-Butyl-cis-3-oxabicyclo[4.3.0]nonan-2-one (3) and trans-4-butyl-cis-3-oxabicyclo[4.3.0]non-7-en-2-one (7) exhibited high fungistatic activity (IC50 = 143.2 and 110.2 μg/mL, respectively) against P. citrinum AM354 as well. PMID:25110806

  2. [Activity of cefpodoxime and other oral beta-lactams against Haemophilus influenzae and Streptococcus pneumoniae with different susceptibilities to penicillin].

    PubMed

    Fenoll, A; Robledo, O; Lerma, M; Giménez, M J; Cebrián, L; Casal, J; Aguilar, L; Gómez-Lus, M L

    2006-03-01

    This study explores the influence on the intrinsic activity of different oral beta-lactams of beta-lactamase production in Haemophilus influenzae and penicillin resistance in Streptococcus pneumoniae. Three substudies were performed: a) a general susceptibility study, analyzing 550 strains received by the Spanish Laboratorio de Referencia de Neumococos throughout February and March 2005; b) a study on the influence of penicillin resistance on the activity of beta-lactams, analyzing 251 penicillin-susceptible strains (MICor=2 mg/l) randomly chosen among those received by the Spanish Laboratorio de Referencia de Neumococos throughout 2005; and c) an H. influenzae susceptibility study analyzing 150 strains received by Instituto Valenciano de Microbiologia throughout 2005. A total of 71% of S. pneumoniae strains were susceptible to penicillin, 21% exhibited intermediate resistance and 8% strains presented full resistance. H. influenzae beta-lactamase production rate was 18.6%. Of the non-beta-lactamase-producing strains, 3% were not susceptible to ampicillin. Cefpodoxime and cefixime exhibited the highest intrinsic activity against H. influenzae, while amoxicillin and cefpodoxime were the most active compounds against S. pneumoniae. All H. influenzae strains were susceptible to oral cephalosporins and amoxicillin/clavulanic acid. The increase in penicillin resistance in S. pneumoniae influenced cefixime, cefaclor and cefuroxime to a higher degree than amoxicillin and cefpodoxime.

  3. Resistance to β-lactam antibiotic may influence nanH gene expression in Trueperella pyogenes isolated from bovine endometritis.

    PubMed

    Zhang, De-Xian; Tian, Kai; Han, Li-Mei; Wang, Qiu-Xia; Liu, Yao-Chuan; Tian, Chun-Lian; Liu, Ming-Chun

    2014-01-01

    Virulence could be modulated by many instinctive and environmental factors including oxygen, osmolarity and antimicrobial agents. This study aimed to investigate the correlation between drug resistance and the nanH expression in Trueperella pyogenes (T. pyogenes). Minimum inhibitory concentrations (MICs) of 6 β-lactam antimicrobial agents (penicillin G, amoxicillin, oxacillin, cefazolin, ceftiofur, and ampicillin) against T. pyogenes were tested by standard broth dilution method according to the protocols of the Clinical and Laboratory Standards Institute (CLSI), and real-time fluorescent quantitative reverse transcription-polymerase chain reaction (RT-PCR) was selected to investigate the mRNA expression levels of the nanH in T. pyogenes. All the isolates were resistant to atleast 2 of antimicrobial agents, and multidrug resistance (resistance to atleast 3 antimicrobials) was observed in 84.38% (27/32) of isolates. The mRNA expression levels of the nanH were significantly higher in comparison with that in ATCC19411, as the resistance profile enlarged, the nanH mRNA expression levels decreased in T. pyogenes. These results indicated that β-lactam antibiotic resistance in T. pyogenes may alter the expression of the nanH. PMID:24803199

  4. Hydroxyl-radical-induced degradative oxidation of beta-lactam antibiotics in water: absolute rate constant measurements.

    PubMed

    Dail, Michelle K; Mezyk, Stephen P

    2010-08-19

    The beta-lactam antibiotics are some of the most prevalent pharmaceutical contaminants currently being detected in aquatic environments. Because the presence of any trace level of antibiotic in water may adversely affect aquatic ecosystems and contribute to the production of antibiotic-resistant bacteria, active removal by additional water treatments, such as using advanced oxidation and reduction processes (AO/RPs), may be required. However, to ensure that any AOP treatment process occurs efficiently and quantitatively, a full understanding of the kinetics and mechanisms of all of the chemical reactions involved under the conditions of use is necessary. In this study, we report on our kinetic measurements for the hydroxyl-radical-induced oxidation of 11 beta-lactam antibiotics obtained using electron pulse radiolysis techniques. For the 5-member ring species, an average reaction rate constant of (7.9 +/- 0.8) x 10(9) M(-1) s(-1) was obtained, slightly faster than for the analogous 6-member ring containing antibiotics, (6.6 +/- 1.2) x 10(9) M(-1) s(-1). The consistency of these rate constants for each group infers a common reaction mechanism, consisting of the partitioning of the hydroxyl radical between addition to peripheral aromatic rings and reaction with the central double-ring core of these antibiotics.

  5. Appearance of β-lactam Resistance Genes in Agricultural Soils and Clinical Isolates over the 20th Century

    PubMed Central

    Graham, David W.; Knapp, Charles W.; Christensen, Bent T.; McCluskey, Seánín; Dolfing, Jan

    2016-01-01

    Debate exists about whether agricultural versus medical antibiotic use drives increasing antibiotic resistance (AR) across nature. Both sectors have been inconsistent at antibiotic stewardship, but it is unclear which sector has most influenced acquired AR on broad scales. Using qPCR and soils archived since 1923 at Askov Experimental Station in Denmark, we quantified four broad-spectrum β-lactam AR genes (ARG; blaTEM, blaSHV, blaOXA and blaCTX-M) and class-1 integron genes (int1) in soils from manured (M) versus inorganic fertilised (IF) fields. “Total” β-lactam ARG levels were significantly higher in M versus IF in soils post-1940 (paired-t test; p < 0.001). However, dominant individual ARGs varied over time; blaTEM and blaSHV between 1963 and 1974, blaOXA slightly later, and blaCTX-M since 1988. These dates roughly parallel first reporting of these genes in clinical isolates, suggesting ARGs in animal manure and humans are historically interconnected. Archive data further show when non-therapeutic antibiotic use was banned in Denmark, blaCTX-M levels declined in M soils, suggesting accumulated soil ARGs can be reduced by prudent antibiotic stewardship. Conversely, int1 levels have continued to increase in M soils since 1990, implying direct manure application to soils should be scrutinized as part of future stewardship programs. PMID:26878889

  6. Continuous infusion of antibiotics in the critically ill: The new holy grail for beta-lactams and vancomycin?

    PubMed Central

    2012-01-01

    The alarming global rise of antimicrobial resistance combined with the lack of new antimicrobial agents has led to a renewed interest in optimization of our current antibiotics. Continuous infusion (CI) of time-dependent antibiotics has certain theoretical advantages toward efficacy based on pharmacokinetic/pharmacodynamic principles. We reviewed the available clinical studies concerning continuous infusion of beta-lactam antibiotics and vancomycin in critically ill patients. We conclude that CI of beta-lactam antibiotics is not necessarily more advantageous for all patients. Continuous infusion is only likely to have clinical benefits in subpopulations of patients where intermittent infusion is unable to achieve an adequate time above the minimal inhibitory concentration (T > MIC). For example, in patients with infections caused by organisms with elevated MICs, patients with altered pharmacokinetics (such as the critically ill) and possibly also immunocompromised patients. For vancomycin CI can be chosen, not always for better clinical efficacy, but because it is practical, cheaper, associated with less AUC24h (area under the curve >24 h)-variability, and easier to monitor. PMID:22747633

  7. Synergistic effect of (+)-pinitol from Saraca asoca with β-lactam antibiotics and studies on the in silico possible mechanism.

    PubMed

    Ahmad, Furkan; Misra, Laxminarain; Gupta, Vivek Kumar; Darokar, Mahendra Pandurang; Prakash, Om; Khan, Feroz; Shukla, Rakesh

    2016-01-01

    Saraca asoca bark has been used in the Ayurvedic system of medicine for female urino-genital disorders. We have recently reported the isolation and characterization of several compounds as markers to develop HPLC profiling of its methanol and aqueous methanol extracts. Now, a HPLC-PDA inactive compound, (+)-pinitol has been isolated and characterized from the bark of this medicinally important tree. Pinitol is a well known bioactive compound for a variety of biological activities, including hypoglycemic and anti-inflammatory activities. A process for the isolation of relatively good concentration of (+)-pinitol from S. asoca bark has been developed and its in vitro anti TNF-α and anti-inflammatory activities against carragenan-induced edema confirmed. While conducting experiments on the possible agonistic activity, it was found that (+)-pinitol showed up to eight fold reduction in the doses of β-lactam antibiotics. The mechanism of its agonistic activity was studied by docking experiments which showed that different conformations of (+)-pinitol and antibiotics were actually in the same binding site with no significant change in the binding energy. These docking simulations, thus predict the possible binding mode of studied compounds and probable reason behind the synergistic effect of (+)-pinitol along with β-lactam antibiotics.

  8. Appearance of β-lactam Resistance Genes in Agricultural Soils and Clinical Isolates over the 20th Century.

    PubMed

    Graham, David W; Knapp, Charles W; Christensen, Bent T; McCluskey, Seánín; Dolfing, Jan

    2016-01-01

    Debate exists about whether agricultural versus medical antibiotic use drives increasing antibiotic resistance (AR) across nature. Both sectors have been inconsistent at antibiotic stewardship, but it is unclear which sector has most influenced acquired AR on broad scales. Using qPCR and soils archived since 1923 at Askov Experimental Station in Denmark, we quantified four broad-spectrum β-lactam AR genes (ARG; bla(TEM), bla(SHV), bla(OXA) and bla(CTX-M)) and class-1 integron genes (int1) in soils from manured (M) versus inorganic fertilised (IF) fields. "Total" β-lactam ARG levels were significantly higher in M versus IF in soils post-1940 (paired-t test; p < 0.001). However, dominant individual ARGs varied over time; bla(TEM) and bla(SHV) between 1963 and 1974, bla(OXA) slightly later, and bla(CTX-M) since 1988. These dates roughly parallel first reporting of these genes in clinical isolates, suggesting ARGs in animal manure and humans are historically interconnected. Archive data further show when non-therapeutic antibiotic use was banned in Denmark, bla(CTX-M) levels declined in M soils, suggesting accumulated soil ARGs can be reduced by prudent antibiotic stewardship. Conversely, int1 levels have continued to increase in M soils since 1990, implying direct manure application to soils should be scrutinized as part of future stewardship programs. PMID:26878889

  9. The Streptococcus pneumoniae pezAT Toxin–Antitoxin System Reduces β-Lactam Resistance and Genetic Competence

    PubMed Central

    Chan, Wai T.; Espinosa, Manuel

    2016-01-01

    Chromosomally encoded Type II Toxin–Antitoxin operons are ubiquitous in bacteria and archaea. Antitoxins neutralize the toxic effect of cognate Toxins by protein–protein interactions and sequestering the active residues of the Toxin. Toxins target essential bacterial processes, mostly translation and replication. However, one class apart is constituted by the PezAT pair because the PezT toxin target cell wall biosynthesis. Here, we have examined the role of the pezAT toxin–antitoxin genes in its natural host, the pathogenic bacterium Streptococcus pneumoniae. The pezAT operon on Pneumococcal Pathogenicity Island 1 was deleted from strain R6 and its phenotypic traits were compared with those of the wild type. The mutant cells formed shorter chains during exponential phase, leading to increased colony-forming units. At stationary phase, the mutant was more resilient to lysis. Importantly, the mutant exhibited higher resistance to antibiotics targeting cell walls (β-lactams), but not to antibiotics acting at other levels. In addition, the mutants also showed enhanced genetic competence. We suggest that PezAT participates in a subtle equilibrium between loss of functions (resistance to β-lactams and genetic competence) and gain of other traits (virulence). PMID:27610103

  10. Two rhodamine lactam modulated lysosome-targetable fluorescence probes for sensitively and selectively monitoring subcellular organelle pH change.

    PubMed

    Li, Hongmei; Wang, Cuiling; She, Mengyao; Zhu, Yuelu; Zhang, Jidong; Yang, Zheng; Liu, Ping; Wang, Yaoyu; Li, Jianli

    2015-11-01

    Be a powerful technique for convenient detection of pH change in living cells, especially at subcellular level, fluorescent probes has attracted more and more attention. In this work, we designed and synthesized three rhodamine lactam modulated fluorescent probes RS1, RS2 and RS3, which all respond sensitively toward weak acidity (pH range 4-6) via the photophysical property in buffer solution without interference from the other metal ions, and they also show ideal pKa values and excellent reversibility. Particularly, by changing the lone pair electrons distribution of lactam-N atom with different conjugations, RS2 and RS3 exhibit high quantum yield, negligible cytotoxicity and excellent permeability. They are suitable to stain selectively lysosomes of tumor cells and monitor its pH changes sensitively via optical molecular imaging. The above findings suggest that the probes we designed could act as ideal and easy method for investigating the pivotal role of H(+) in lysosomes and are potential pH detectors in disease diagnosis through direct intracellular imaging.

  11. The Streptococcus pneumoniae pezAT Toxin–Antitoxin System Reduces β-Lactam Resistance and Genetic Competence

    PubMed Central

    Chan, Wai T.; Espinosa, Manuel

    2016-01-01

    Chromosomally encoded Type II Toxin–Antitoxin operons are ubiquitous in bacteria and archaea. Antitoxins neutralize the toxic effect of cognate Toxins by protein–protein interactions and sequestering the active residues of the Toxin. Toxins target essential bacterial processes, mostly translation and replication. However, one class apart is constituted by the PezAT pair because the PezT toxin target cell wall biosynthesis. Here, we have examined the role of the pezAT toxin–antitoxin genes in its natural host, the pathogenic bacterium Streptococcus pneumoniae. The pezAT operon on Pneumococcal Pathogenicity Island 1 was deleted from strain R6 and its phenotypic traits were compared with those of the wild type. The mutant cells formed shorter chains during exponential phase, leading to increased colony-forming units. At stationary phase, the mutant was more resilient to lysis. Importantly, the mutant exhibited higher resistance to antibiotics targeting cell walls (β-lactams), but not to antibiotics acting at other levels. In addition, the mutants also showed enhanced genetic competence. We suggest that PezAT participates in a subtle equilibrium between loss of functions (resistance to β-lactams and genetic competence) and gain of other traits (virulence).

  12. In vitro Potential Effect of Morin in the Combination with β-Lactam Antibiotics Against Methicillin-Resistant Staphylococcus aureus.

    PubMed

    Mun, Su-Hyun; Lee, Young-Seob; Han, Sin-Hee; Lee, Sang-Won; Cha, Seon-Woo; Kim, Sung-bae; Seo, Yun-Soo; Kong, Ryong; Kang, Da-Hye; Shin, Dong-Won; Kang, Ok-Hwa; Kwon, Dong-Yeul

    2015-06-01

    Morin, a plant-derived flavonol, is known to be an effective inhibitor of Gram-positive bacteria. In this study, we explored the combined effect of morin with β-lactam antibiotics against methicillin-resistant Staphylococcus aureus (MRSA), a multidrug-resistant pathogen. The anti-MRSA activity of morin was investigated by the broth microdilution method, checkerboard dilution test, and time-kill curve assay. The expression of the resistant protein, penicillin-binding protein (PBP2a) encoded by mecA, was analyzed by the Western blotting method in the presence of morin and oxacillin. An increased susceptibility of MRSA toward oxacillin was observed in the presence of morin. The protein level of PBP2a was reduced when MRSA (ATCC 33591) was treated with the combination of morin and oxacillin, indicating that the combination of morin and oxacillin potentiates the killing effect against MRSA. The present study indicates that the killing effect by the combinative treatment of morin and β-lactam antibiotic is dependent on the PBP2a-mediated resistance mechanism.

  13. In vitro activity of beta-lactam antibiotics to community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA).

    PubMed

    Germel, C; Haag, A; Söderquist, B

    2012-04-01

    Community-associated (CA) MRSA often display low MIC values against oxacillin. The in vitro activity of various beta-lactam antibiotics against heterogeneous CA-MRSA (n = 98) isolated in a low endemic area was determined by Etest, and Mueller-Hinton agar (MUHAP) was compared with Mueller-Hinton agar supplemented with 2% NaCl (MUHSP). In general, the CA-MRSA isolates showed higher MIC values for the various beta-lactam antibiotics on MUHSP compared with MUHAP. MIC values for oxacillin ranged from 1 to >256 mg/L on MUHSP. Cephalothin, representing the first generation of cephalosporins, showed MICs from 0.75 to 96 mg/L and the MIC(50) and MIC(90) for cefuroxime, cefotaxime and cefepime, representing the second, third and fourth generations, respectively, were rather high. However, the MIC(50) and MIC(90) for ceftobiprole (fifth generation) were 1.5 and 2 mg/L, respectively, on MUHSP. The MIC(50) and MIC(90) for imipenem were 0.75 and 2 mg/L, respectively, on MUHSP. Only 3/98 (3%) CA-MRSA isolates showed a MIC >4 mg/L. Consequently, low MIC values for imipenem, lower than those of the newly developed fifth generation cephalosporins, were found among CA-MRSA. These findings may be considered for further studies including clinical trials in order to evaluate carbapenems as a potential treatment option for infections caused by CA-MRSA.

  14. Prevalence of β-lactam (blaTEM) and Metronidazole (nim) Resistance Genes in the Oral Cavity of Greek Subjects

    PubMed Central

    Koukos, Georgios; Konstantinidis, Antonios; Tsalikis, Lazaros; Arsenakis, Minas; Slini, Theodora; Sakellari, Dimitra

    2016-01-01

    Objectives: The aim of this study is to investigate the prevalence of blaTEM and nim genes that encode resistance to β-lactams and nitroimidazoles, respectively, in the oral cavity of systemically healthy Greek subjects. Materials and Methodology: After screening 720 potentially eligible subjects, 154 subjects were recruited for the study, including 50 periodontally healthy patients, 52 cases of gingivitis and 52 cases of chronic periodontitis. The clinical parameters were assessed with an automated probe. Various samples were collected from the tongue, first molars and pockets >6mm, and analysed by polymerase chain reaction-amplification of the blaTEM and nim genes, using primers and conditions previously described in the literature. Results: There was a high rate of detection of blaTEM in plaque and tongue samples alike in all periodontal conditions (37% of plaque and 60% of tongue samples, and 71% of participants). The blaTEM gene was detected more frequently in the tongue samples of the periodontally healthy (56%) and chronic periodontitis (62%) groups compared to the plaque samples from the same groups (36% and 29%, respectively; z-test with Bonferroni corrections-tests, P<0.05). The nim gene was not detected in any of the 343 samples analysed. Conclusion: The oral cavity of Greek subjects often harbours blaTEM but not nim genes, and therefore the antimicrobial activity of β-lactams might be compromised. PMID:27099637

  15. The Streptococcus pneumoniae pezAT Toxin-Antitoxin System Reduces β-Lactam Resistance and Genetic Competence.

    PubMed

    Chan, Wai T; Espinosa, Manuel

    2016-01-01

    Chromosomally encoded Type II Toxin-Antitoxin operons are ubiquitous in bacteria and archaea. Antitoxins neutralize the toxic effect of cognate Toxins by protein-protein interactions and sequestering the active residues of the Toxin. Toxins target essential bacterial processes, mostly translation and replication. However, one class apart is constituted by the PezAT pair because the PezT toxin target cell wall biosynthesis. Here, we have examined the role of the pezAT toxin-antitoxin genes in its natural host, the pathogenic bacterium Streptococcus pneumoniae. The pezAT operon on Pneumococcal Pathogenicity Island 1 was deleted from strain R6 and its phenotypic traits were compared with those of the wild type. The mutant cells formed shorter chains during exponential phase, leading to increased colony-forming units. At stationary phase, the mutant was more resilient to lysis. Importantly, the mutant exhibited higher resistance to antibiotics targeting cell walls (β-lactams), but not to antibiotics acting at other levels. In addition, the mutants also showed enhanced genetic competence. We suggest that PezAT participates in a subtle equilibrium between loss of functions (resistance to β-lactams and genetic competence) and gain of other traits (virulence). PMID:27610103

  16. Oral Gram-negative anaerobic bacilli as a reservoir of β-lactam resistance genes facilitating infections with multiresistant bacteria.

    PubMed

    Dupin, Clarisse; Tamanai-Shacoori, Zohreh; Ehrmann, Elodie; Dupont, Anais; Barloy-Hubler, Frédérique; Bousarghin, Latifa; Bonnaure-Mallet, Martine; Jolivet-Gougeon, Anne

    2015-02-01

    Many β-lactamases have been described in various Gram-negative bacilli (Capnocytophaga, Prevotella, Fusobacterium, etc.) of the oral cavity, belonging to class A of the Ambler classification (CepA, CblA, CfxA, CSP-1 and TEM), class B (CfiA) or class D in Fusobacterium nucleatum (FUS-1). The minimum inhibitory concentrations of β-lactams are variable and this variation is often related to the presence of plasmids or other mobile genetic elements (MGEs) that modulate the expression of resistance genes. DNA persistence and bacterial promiscuity in oral biofilms also contribute to genetic transformation and conjugation in this particular microcosm. Overexpression of efflux pumps is facilitated because the encoding genes are located on MGEs, in some multidrug-resistant clinical isolates, similar to conjugative transposons harbouring genes encoding β-lactamases. All these facts lead us to consider the oral cavity as an important reservoir of β-lactam resistance genes and a privileged place for genetic exchange, especially in commensal strictly anaerobic Gram-negative bacilli.

  17. Hybrid anticancer compounds. Steroidal lactam esters of carboxylic derivatives of N,N-bis (2-chloroethyl) aniline (review).

    PubMed

    Catsoulacos, P; Catsoulacos, D

    1991-01-01

    For the rational design of more specific alkylating agents, we suggested new biological platforms able to deliver the alkylating moieties to specific target site and on the other hand we hoped to lead in compounds with synergistic activity. As biological platforms have been used steroidal lactams of A and D- ring and as alkylating agents carboxylic derivatives of N,N-bis (2-Chloroethyl) aniline which combine to the steroid by an easily cleaved ester bond. These homo-aza-steroidal esters gave satisfactory results in early and advanced P388, L1210 leukemias and solid tumors. Whereas unmodified steroidal esters have generally been reported to be inactive in treatment of L1210 leukemia. The steric arrangement of the alkylating moiety greatly effects toxicity and activity of the drugs, while the steric arrangement of the hydrogen atom at position 5 influences these parameters. Isosterism of alkylating agent is the factor for biological action. The amide group of the lactam molecule may be essential for activity.

  18. General approach for the stereocontrolled construction of the beta-lactam ring in amino acid-derived 4-alkyl-4-carboxy-2-azetidinones.

    PubMed

    Gerona-Navarro, Guillermo; García-López, M Teresa; González-Muñiz, Rosario

    2002-05-31

    The first general approach toward the asymmetric synthesis of 4-alkyl-4-carboxy-2-azetidinones derived from amino acids is described. The stereoselective construction of the beta-lactam ring was achieved through base-mediated intramolecular cyclization of the corresponding N(alpha)-chloroacetyl derivatives bearing (+)- or (-)-10-(N,N-dicyclohexylsulfamoyl)isoborneol as chiral auxiliary (ee up to 82%).

  19. Pyrrolidine-5,5-trans-lactams as novel mechanism-based inhibitors of human cytomegalovirus protease. Part 3: potency and plasma stability.

    PubMed

    Borthwick, Alan D; Exall, Anne M; Haley, Terry M; Jackson, Deborah L; Mason, Andrew M; Weingarten, Gordon G

    2002-07-01

    Mechanism-based inhibitors of HCMV protease, which are stable to human plasma (> or = 20 h) and have single-figure potency in the microM range against HCMV protease, have been developed based on the dansylproline alpha-methyl pyrrolidine-5,5-trans-lactam nucleus.

  20. Syntheses and studies of new forms of N-sulfonyloxy β-lactams as potential antibacterial agents and β-lactamase inhibitors.

    PubMed

    Carosso, Serena; Miller, Marvin J

    2015-09-15

    The synthesis of a small library of N-sulfonyloxy-2-azetidinones is reported and the preliminary results of the investigation of the biological activity of these molecules are discussed. These new multi-electrophilic β-lactams ('electrophilic bombs') display unexpected selectivity in their antibacterial activity and β-lactamase inhibitory activity. PMID:26321604

  1. Syntheses and studies of new forms of N-sulfonyloxy β-lactams as potential antibacterial agents and β-lactamase inhibitors.

    PubMed

    Carosso, Serena; Miller, Marvin J

    2015-09-15

    The synthesis of a small library of N-sulfonyloxy-2-azetidinones is reported and the preliminary results of the investigation of the biological activity of these molecules are discussed. These new multi-electrophilic β-lactams ('electrophilic bombs') display unexpected selectivity in their antibacterial activity and β-lactamase inhibitory activity.

  2. Syntheses and Studies of New Forms of N-Sulfonyloxy β-Lactams as Potential Antibacterial Agents and β-Lactamase Inhibitors

    PubMed Central

    Carosso, Serena; Miller, Marvin J.

    2015-01-01

    The synthesis of a small library of N-sulfonyloxy-2-azetidinones is reported and the preliminary results of the investigation of the biological activity of these molecules are discussed. These new multi-electrophilic β-lactams (“electrophilic bombs”) display unexpected selectivity in their antibacterial activity and β-lactamase inhibitory activity. PMID:26321604

  3. One-pot preparation of trifluoromethylated homoallylic N-acylhydrazines or α-methylene-γ-lactams from acylhydrazines, trifluoroacetaldehyde methyl hemiacetal, allyl bromide and tin.

    PubMed

    Du, Ganggang; Huang, Danfeng; Wang, Ke-Hu; Chen, Xiaowei; Xu, Yanli; Ma, Junyan; Su, Yingpeng; Fu, Ying; Hu, Yulai

    2016-01-28

    An efficient and convenient one-pot method for the preparation of trifluoromethylated homoallylic N-acylhydrazines or α-methylene-γ-lactams has been described. In this process, allyl bromide and metal tin are used instead of toxic stannanes, and commercially available aqueous trifluoroacetaldehyde methyl hemiacetal was used as a trifluoromethyl source.

  4. Synthesis of dibenzoxepine lactams via a Cu-catalyzed one-pot etherification/aldol condensation cascade reaction: application toward the total synthesis of aristoyagonine.

    PubMed

    Lim, Hye Sun; Choi, Young Lok; Heo, Jung-Nyoung

    2013-09-20

    A general synthesis of dibenzoxepine lactams has been developed using a one-pot Cu-catalyzed etherification/aldol condensation cascade reaction. The reaction of 4-hydroxyisoindolin-1-one with a wide range of 2-bromobenzaldehydes in the presence of a copper catalyst provided various aristoyagonine derivatives in good yields. PMID:24000941

  5. Development of test panel of beta-lactamases expressed in a common Escherichia coli host background for evaluation of new beta-lactam antibiotics.

    PubMed Central

    Bradford, P A; Sanders, C C

    1995-01-01

    A test panel of 35 different beta-lactamases expressed in a common Escherichia coli host was created to compare the effect that each beta-lactamase had on susceptibility to various beta-lactam antibiotics. A comparison of the MICs obtained with this panel generally reflected differences in the substrate profiles of the various beta-lactamases examined. In addition, several strains of the panel were subjected to selection with porin-specific bacteriophages to obtain mutants lacking either the OmpC or OmpF porin protein. A mutation in either OmpC or OmpF did change the susceptibilities of certain strains expressing beta-lactamase to certain beta-lactam antibiotics. However, the loss of a single porin did not predictably alter susceptibility to any given beta-lactam drug. This panel of strains producing various beta-lactamases was found to be a useful tool for comparing the effects of different beta-lactamases and outer membrane permeability upon susceptibility to beta-lactam drugs. PMID:7726487

  6. Discovery of substituted lactams as novel dual orexin receptor antagonists. Synthesis, preliminary structure-activity relationship studies and efforts towards improved metabolic stability and pharmacokinetic properties. Part 1.

    PubMed

    Sifferlen, Thierry; Boller, Amandine; Chardonneau, Audrey; Cottreel, Emmanuelle; Hoecker, Johannes; Aissaoui, Hamed; Williams, Jodi T; Brotschi, Christine; Heidmann, Bibia; Siegrist, Romain; Gatfield, John; Treiber, Alexander; Brisbare-Roch, Catherine; Jenck, Francois; Boss, Christoph

    2014-02-15

    Starting from a thiazolidin-4-one HTS hit, a novel series of substituted lactams was identified and developed as dual orexin receptor antagonists. In this Letter, we describe our initial efforts towards the improvement of potency and metabolic stability. These investigations delivered optimized lead compounds with CNS drug-like properties suitable for further optimization. PMID:24447850

  7. Restoration of susceptibility of intracellular methicillin-resistant Staphylococcus aureus to beta-lactams: comparison of strains, cells, and antibiotics.

    PubMed

    Lemaire, Sandrine; Olivier, Aurélie; Van Bambeke, Françoise; Tulkens, Paul M; Appelbaum, Peter C; Glupczynski, Youri

    2008-08-01

    Staphylococcus aureus invades eukaryotic cells. When methicillin-resistant S. aureus (MRSA) ATCC 33591 is phagocytized by human THP-1 macrophages, complete restoration of susceptibility to cloxacillin and meropenem is shown and the strain becomes indistinguishable from MSSA ATCC 25923 due to the acid pH prevailing in phagolysosomes (S. Lemaire et al., Antimicrob. Agents Chemother. 51:1627-1632, 2007). We examined whether this observation can be extended to (i) strains of current clinical and epidemiological interest (three hospital-acquired MRSA [HA-MRSA] strains, two community-acquired MRSA [CA-MRSA] strains, two HA-MRSA strains with the vancomycin-intermediate phenotype, one HA-MRSA strain with the vancomycin-resistant phenotype, and one animal [porcine] MRSA strain), (ii) activated THP-1 cells and nonprofessional phagocytes (keratinocytes, Calu-3 bronchial epithelial cells), and (iii) other beta-lactams (imipenem, oxacillin, cefuroxime, cefepime). All strains showed (i) a marked reduction in MICs in broth at pH 5.5 compared with the MIC at pH 7.4 and (ii) sigmoidal dose-response curves with cloxacillin (0.01x to 100x MIC, 24 h of incubation) after phagocytosis by THP-1 macrophages that were indistinguishable from each other and from the dose-response curve for methicillin-susceptible S. aureus (MSSA) ATCC 25923 (relative potency [50% effect], 6.09x MIC [95% confidence interval {CI}, 4.50 to 8.25]; relative efficacy [change in bacterial counts over the original inoculum for an infinitely large cloxacillin concentration, or maximal effect], -0.69 log CFU [95% CI, -0.79 to -0.58]). Similar dose-response curves for cloxacillin were also observed with MSSA ATCC 25923 and MRSA ATCC 33591 after phagocytosis by activated THP-1 macrophages, keratinocytes, and Calu-3 cells. By contrast, there was a lower level of restoration of susceptibility of MRSA ATCC 33591 to cefuroxime and cefepime after phagocytosis by THP-1 macrophages, even when the data were normalized for

  8. Crystal Structures of Covalent Complexes of [beta]-Lactam Antibiotics with Escherichia coli Penicillin-Binding Protein 5: Toward an Understanding of Antibiotic Specificity

    SciTech Connect

    Nicola, George; Tomberg, Joshua; Pratt, R.F.; Nicholas, Robert A.; Davies, Christopher

    2010-12-07

    Penicillin-binding proteins (PBPs) are the molecular targets for the widely used {beta}-lactam class of antibiotics, but how these compounds act at the molecular level is not fully understood. We have determined crystal structures of Escherichia coli PBP 5 as covalent complexes with imipenem, cloxacillin, and cefoxitin. These antibiotics exhibit very different second-order rates of acylation for the enzyme. In all three structures, there is excellent electron density for the central portion of the {beta}-lactam, but weak or absent density for the R1 or R2 side chains. Areas of contact between the antibiotics and PBP 5 do not correlate with the rates of acylation. The same is true for conformational changes, because although a shift of a loop leading to an electrostatic interaction between Arg248 and the {beta}-lactam carboxylate, which occurs completely with cefoxitin and partially with imipenem and is absent with cloxacillin, is consistent with the different rates of acylation, mutagenesis of Arg248 decreased the level of cefoxitin acylation only 2-fold. Together, these data suggest that structures of postcovalent complexes of PBP 5 are unlikely to be useful vehicles for the design of new covalent inhibitors of PBPs. Finally, superimposition of the imipenem-acylated complex with PBP 5 in complex with a boronic acid peptidomimetic shows that the position corresponding to the hydrolytic water molecule is occluded by the ring nitrogen of the {beta}-lactam. Because the ring nitrogen occupies a similar position in all three complexes, this supports the hypothesis that deacylation is blocked by the continued presence of the leaving group after opening of the {beta}-lactam ring.

  9. β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus USA300 Is Increased by Inactivation of the ClpXP Protease

    PubMed Central

    Bæk, Kristoffer T.; Gründling, Angelika; Mogensen, René G.; Thøgersen, Louise; Petersen, Andreas; Paulander, Wilhelm

    2014-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) has acquired the mecA gene encoding a peptidoglycan transpeptidase, penicillin binding protein 2a (PBP2a), which has decreased affinity for β-lactams. Quickly spreading and highly virulent community-acquired (CA) MRSA strains recently emerged as a frequent cause of infection in individuals without exposure to the health care system. In this study, we found that the inactivation of the components of the ClpXP protease substantially increased the β-lactam resistance level of a CA-MRSA USA300 strain, suggesting that the proteolytic activity of ClpXP controls one or more pathways modulating β-lactam resistance. These pathways do not involve the control of mecA expression, as the cellular levels of PBP2a were unaltered in the clp mutants. An analysis of the cell envelope properties of the clpX and clpP mutants revealed a number of distinct phenotypes that may contribute to the enhanced β-lactam tolerance. Both mutants displayed significantly thicker cell walls, increased peptidoglycan cross-linking, and altered composition of monomeric muropeptide species compared to those of the wild types. Moreover, changes in Sle1-mediated peptidoglycan hydrolysis and altered processing of the major autolysin Atl were observed in the clp mutants. In conclusion, the results presented here point to an important role for the ClpXP protease in controlling cell wall metabolism and add novel insights into the molecular factors that determine strain-dependent β-lactam resistance. PMID:24867990

  10. β-Lactam resistance in methicillin-resistant Staphylococcus aureus USA300 is increased by inactivation of the ClpXP protease.

    PubMed

    Bæk, Kristoffer T; Gründling, Angelika; Mogensen, René G; Thøgersen, Louise; Petersen, Andreas; Paulander, Wilhelm; Frees, Dorte

    2014-08-01

    Methicillin-resistant Staphylococcus aureus (MRSA) has acquired the mecA gene encoding a peptidoglycan transpeptidase, penicillin binding protein 2a (PBP2a), which has decreased affinity for β-lactams. Quickly spreading and highly virulent community-acquired (CA) MRSA strains recently emerged as a frequent cause of infection in individuals without exposure to the health care system. In this study, we found that the inactivation of the components of the ClpXP protease substantially increased the β-lactam resistance level of a CA-MRSA USA300 strain, suggesting that the proteolytic activity of ClpXP controls one or more pathways modulating β-lactam resistance. These pathways do not involve the control of mecA expression, as the cellular levels of PBP2a were unaltered in the clp mutants. An analysis of the cell envelope properties of the clpX and clpP mutants revealed a number of distinct phenotypes that may contribute to the enhanced β-lactam tolerance. Both mutants displayed significantly thicker cell walls, increased peptidoglycan cross-linking, and altered composition of monomeric muropeptide species compared to those of the wild types. Moreover, changes in Sle1-mediated peptidoglycan hydrolysis and altered processing of the major autolysin Atl were observed in the clp mutants. In conclusion, the results presented here point to an important role for the ClpXP protease in controlling cell wall metabolism and add novel insights into the molecular factors that determine strain-dependent β-lactam resistance.

  11. Stereoselective synthesis of densely functionalized pyrrolidin-2-ones by a conjugate addition/nitro-Mannich/lactamization reaction.

    PubMed

    Anderson, James C; Horsfall, Lisa R; Kalogirou, Andreas S; Mills, Matthew R; Stepney, Gregory J; Tizzard, Graham J

    2012-07-20

    Copper-catalyzed conjugate addition of diorgano zinc reagents to nitroacrylate 1 followed by a subsequent nitro-Mannich reaction and in situ lactamization leads to an efficient one-pot synthesis of 1,3,5-trisubstituted 4-nitropyrrolidin-2-ones (5). The versatility of the reaction is shown for a wide range of N-p-(methoxy)phenyl protected aldimines 3 derived from alkyl, aryl, and heteroaryl aldehydes. The densely functionalized pyrrolidin-2-ones 5 are isolated as single diastereoisomers (40 examples, 33-84% yield). An enantioselective copper-catalyzed conjugate addition of diethylzinc led to highly crystalline products that could be recrystallized to enantiopurity in high yield. A range of successful chemoselective transformations were investigated, which widens the applicability of the pyrrolidn-2-ones as stereochemically pure building blocks for further organic synthesis. PMID:22708711

  12. [Is monotherapy with β-lactam antibiotics still up to date? New aspects for treatment of severe infections].

    PubMed

    Heizmann, P; Lode, H; Heizmann, W R

    2012-02-01

    Mortality of sepsis is still high. Crucial for therapeutic response are the early start of treatment as well as the choice of antibiotics or antibiotic combinations. β-lactam antibiotics with bactericidal mode of action are often recommended in guidelines. But this antibiotic class can trigger the immune system to a maximum by releasing cell wall components or exotoxins. This may lead to a worsening of the patient's clinical situation. In contrast, antibiotics with bacteriostatic action often inhibit bacterial protein synthesis with decrease of production of virulence factors and minimize release of cell wall components. The purpose of this review is to summarise the significance of some bacteriostatic antibiotics and to discuss whether a combination of bactericidal and bacteriostatic agents may improve the course of the illness.

  13. Anticancer drug design using scaffolds of β-lactams, sulfonamides, quinoline, quinoxaline and natural products. Drugs advances in clinical trials.

    PubMed

    Balderas-Renteria, I; Gonzalez-Barranco, P; Garcia, A; Banik, B K; Rivera, G

    2012-01-01

    Eleven years after the start of a new millennium characterized by amazing scientific development, the cure for cancer remains a major challenge for humanity. In this regard, scientific efforts have focused on the search for new therapeutic targets that involve specific recognition and stop the spread of cancer cells, as well as the development of new therapeutic options that show greater specificity and better therapeutic efficacy. This review includes recent published literature about new anticancer drug design using scaffolds of β-lactams, sulfonamides, quinoline, quinoxaline and natural products, and focuses on the structure-activity relationships of scaffolds that have been reported to potently inhibit cell growth of human tumor cell lines. It describes not only those synthetic or natural compounds aimed at specific molecular targets of cancer cells in vitro, but also compounds currently in clinical trials.

  14. Pharmacokinetic Drug Interactions of Antimicrobial Drugs: A Systematic Review on Oxazolidinones, Rifamycines, Macrolides, Fluoroquinolones, and Beta-Lactams

    PubMed Central

    Bolhuis, Mathieu S.; Panday, Prashant N.; Pranger, Arianna D.; Kosterink, Jos G. W.; Alffenaar, Jan-Willem C.

    2011-01-01

    Like any other drug, antimicrobial drugs are prone to pharmacokinetic drug interactions. These drug interactions are a major concern in clinical practice as they may have an effect on efficacy and toxicity. This article provides an overview of all published pharmacokinetic studies on drug interactions of the commonly prescribed antimicrobial drugs oxazolidinones, rifamycines, macrolides, fluoroquinolones, and beta-lactams, focusing on systematic research. We describe drug-food and drug-drug interaction studies in humans, affecting antimicrobial drugs as well as concomitantly administered drugs. Since knowledge about mechanisms is of paramount importance for adequate management of drug interactions, the most plausible underlying mechanism of the drug interaction is provided when available. This overview can be used in daily practice to support the management of pharmacokinetic drug interactions of antimicrobial drugs. PMID:24309312

  15. Neisseria lactamica and Neisseria polysaccharea as possible sources of meningococcal beta-lactam resistance by genetic transformation.

    PubMed Central

    Saez-Nieto, J A; Lujan, R; Martinez-Suarez, J V; Berron, S; Vazquez, J A; Viñas, M; Campos, J

    1990-01-01

    We studied the susceptibilities of relatively penicillin G-resistant and -susceptible strains of Neisseria meningitidis, as well as Neisseria lactamica and Neisseria polysaccharea, to penicillin, ampicillin, and several cephalosporins. The MICs of penicillin, ampicillin, cephalothin, and cefuroxime for moderately resistant meningococci have increased two- to sixfold in relation to MICs for susceptible strains. For these strains of meningococci, N. lactamica, and N. polysaccharea, penicillin, ampicillin, cephalothin, and cefuroxime MICs for 50 and 90% of strains were similar. By genetic transformation of a penicillin-susceptible strain of N. meningitidis to low-level penicillin resistance with DNA from penicillin-resistant strains of N. meningitidis, N. lactamica, N. polysaccharea, and N. gonorrhoeae, isogenic strains with the same pattern of resistance to beta-lactams were obtained, suggesting that these commensal Neisseria spp. could be the source of meningococcal resistance genes. PMID:2127349

  16. Lipase mediated chiral resolution of 4-arylthio-2-butanol as an intermediate for beta-lactam antibiotics.

    PubMed

    Hwang, Kwang-Jin; Lee, Jinkue; Chin, Sungmin; Moon, Chi Jang; Lee, Wonjae; Baek, Chae-Sun; Kim, Hyung Jin

    2003-12-01

    This paper deals with chiral enzymatic resolution of 4-arylthio-2-butanols by lipase to prepare potential intermediates of beta-lactam antibiotics. Among several lipases employed, lipase P type enzyme gave the highest ee value to prepare (R)-4-arylthio-2-butyl acetate. The enzymatic resolution of phenyl substituted alcohol (6a) using lipase P showed the highest ee value (99.7%) among those of 4-arylthio-2-butanol derivatives. Lipase P mediated hydrolysis of acylester 7a gave also (R)-alcohol 6a selectively. For determination of enantiomeric purity of these enzymatic resolved analytes, liquid chromatographic analysis was performed using two coupled Chiralcel OD and (R,R)-WhelkO chiral column.

  17. An analysis of beta-lactam-derived antigens on spleen cell and serum proteins by ELISA and Western blotting.

    PubMed

    Warbrick, E V; Thomas, A L; Stejskal, V; Coleman, J W

    1995-11-01

    Penicillins and related beta-lactam antibiotics are known to conjugate to proteins to generate potentially antigenic (haptenic) determinants. In the present study, we used a rabbit polyclonal antibody raised against benzylpenicillin (BP) to investigate the capacity of six penicillins and one cephalosporin to generate haptenic groups in vitro on cultured mouse spleen cells and on serum proteins in the culture medium. All of the drugs tested, namely, BP, amoxicillin (AMX), ampicillin (AMP), cephalothin (CEP), cloxacillin (CLX), flucloxacillin (FLX), and phenoxymethylpenicillin (PMP) generated antigens in a concentration-dependent manner on cell and serum proteins, which could be detected by ELISA, although antigens generated by BP, CEP, FLX, or PMP in either cell- or serum-conjugated form were more readily detected than those generated by AMX, AMP, or CLX. Western blot analysis revealed that BP-derived antigens were generated relatively slowly on cell proteins (maximum binding was not yet reached after 8 h), compared to serum proteins (maximum binding within 1 h). BP, CEP, and PMP all generated similar distinctive patterns of immunostaining of electrophoresed cell or serum proteins which did not reflect the relative abundance of different proteins as revealed by Coomassie brilliant blue staining. FLX, CLX, AMP, and AMX did not generate antigens that could be detected on Western blots. In conclusion, we have shown that various beta-lactam antibiotics generate antigens on cell and serum proteins that can be detected and characterized immunochemically with polyclonal antiserum. Further application of these methods may offer potential for further identification of immunologically relevant cellular and serum antigens generated by these drugs.

  18. A response regulator from a soil metagenome enhances resistance to the β-lactam antibiotic carbenicillin in Escherichia coli.

    PubMed

    Allen, Heather K; An, Ran; Handelsman, Jo; Moe, Luke A

    2015-01-01

    Functional metagenomic analysis of soil metagenomes is a method for uncovering as-yet unidentified mechanisms for antibiotic resistance. Here we report an unconventional mode by which a response regulator derived from a soil metagenome confers resistance to the β-lactam antibiotic carbenicillin in Escherichia coli. A recombinant clone (βlr16) harboring a 5,169 bp DNA insert was selected from a metagenomic library previously constructed from a remote Alaskan soil. The βlr16 clone conferred specific resistance to carbenicillin, with limited increases in resistance to other tested antibiotics, including other β-lactams (penicillins and cephalosporins), rifampin, ciprofloxacin, erythromycin, chloramphenicol, nalidixic acid, fusidic acid, and gentamicin. Resistance was more pronounced at 24°C than at 37°C. Zone-of-inhibition assays suggested that the mechanism of carbenicillin resistance was not due to antibiotic inactivation. The DNA insert did not encode any genes known to confer antibiotic resistance, but did have two putative open reading frames (ORFs) that were annotated as a metallopeptidase and a two-component response regulator. Transposon mutagenesis and subcloning of the two ORFs followed by phenotypic assays showed that the response regulator gene was necessary and sufficient to confer the resistance phenotype. Quantitative reverse transcriptase PCR showed that the response regulator suppressed expression of the ompF porin gene, independently of the small RNA regulator micF, and enhanced expression of the acrD, mdtA, and mdtB efflux pump genes. This work demonstrates that antibiotic resistance can be achieved by the modulation of gene regulation by heterologous DNA. Functional analyses such as these can be important for making discoveries in antibiotic resistance gene biology and ecology. PMID:25782011

  19. Identification of aminoglycoside and β-lactam resistance genes from within an infant gut functional metagenomic library.

    PubMed

    Fouhy, Fiona; Ogilvie, Lesley A; Jones, Brian V; Ross, R Paul; Ryan, Anthony C; Dempsey, Eugene M; Fitzgerald, Gerald F; Stanton, Catherine; Cotter, Paul D

    2014-01-01

    The infant gut microbiota develops rapidly during the first 2 years of life, acquiring microorganisms from diverse sources. During this time, significant opportunities exist for the infant to acquire antibiotic resistant bacteria, which can become established and constitute the infant gut resistome. With increased antibiotic resistance limiting our ability to treat bacterial infections, investigations into resistance reservoirs are highly pertinent. This study aimed to explore the nascent resistome in antibiotically-naïve infant gut microbiomes, using a combination of metagenomic approaches. Faecal samples from 22 six-month-old infants without previous antibiotic exposure were used to construct a pooled metagenomic library, which was functionally screened for ampicillin and gentamicin resistance. Our library of ∼220Mb contained 0.45 ampicillin resistant hits/Mb and 0.059 gentamicin resistant hits/Mb. PCR-based analysis of fosmid clones and uncloned metagenomic DNA, revealed a diverse and abundant aminoglycoside and β-lactam resistance reservoir within the infant gut, with resistance determinants exhibiting homology to those found in common gut inhabitants, including Escherichia coli, Enterococcus sp., and Clostridium difficile, as well as to genes from cryptic environmental bacteria. Notably, the genes identified differed from those revealed when a sequence-driven PCR-based screen of metagenomic DNA was employed. Carriage of these antibiotic resistance determinants conferred substantial, but varied (2-512x), increases in antibiotic resistance to their bacterial host. These data provide insights into the infant gut resistome, revealing the presence of a varied aminoglycoside and β-lactam resistance reservoir even in the absence of selective pressure, confirming the infant resistome establishes early in life, perhaps even at birth. PMID:25247417

  20. Identification of Aminoglycoside and β-Lactam Resistance Genes from within an Infant Gut Functional Metagenomic Library

    PubMed Central

    Fouhy, Fiona; Ogilvie, Lesley A.; Jones, Brian V.; Ross, R. Paul; Ryan, Anthony C.; Dempsey, Eugene M.; Fitzgerald, Gerald F.; Stanton, Catherine; Cotter, Paul D.

    2014-01-01

    The infant gut microbiota develops rapidly during the first 2 years of life, acquiring microorganisms from diverse sources. During this time, significant opportunities exist for the infant to acquire antibiotic resistant bacteria, which can become established and constitute the infant gut resistome. With increased antibiotic resistance limiting our ability to treat bacterial infections, investigations into resistance reservoirs are highly pertinent. This study aimed to explore the nascent resistome in antibiotically-naïve infant gut microbiomes, using a combination of metagenomic approaches. Faecal samples from 22 six-month-old infants without previous antibiotic exposure were used to construct a pooled metagenomic library, which was functionally screened for ampicillin and gentamicin resistance. Our library of ∼220Mb contained 0.45 ampicillin resistant hits/Mb and 0.059 gentamicin resistant hits/Mb. PCR-based analysis of fosmid clones and uncloned metagenomic DNA, revealed a diverse and abundant aminoglycoside and β-lactam resistance reservoir within the infant gut, with resistance determinants exhibiting homology to those found in common gut inhabitants, including Escherichia coli, Enterococcus sp., and Clostridium difficile, as well as to genes from cryptic environmental bacteria. Notably, the genes identified differed from those revealed when a sequence-driven PCR-based screen of metagenomic DNA was employed. Carriage of these antibiotic resistance determinants conferred substantial, but varied (2–512x), increases in antibiotic resistance to their bacterial host. These data provide insights into the infant gut resistome, revealing the presence of a varied aminoglycoside and β-lactam resistance reservoir even in the absence of selective pressure, confirming the infant resistome establishes early in life, perhaps even at birth. PMID:25247417

  1. Antimicrobial Susceptibilities and Distribution of Resistance Genes for β-Lactams in Streptococcus pneumoniae Isolated in Hamadan

    PubMed Central

    Najafi Mosleh, Mohammad; Gharibi, Marzieh; Alikhani, Mohammad Yousef; Saidijam, Massoud; Kalantarian, Giti

    2014-01-01

    Background: β-lactams resistant Streptococcus pneumoniae are an emerging problem throughout the world. Several resistance mechanisms have been reported, including expression of drug-destroying enzymes such as β-lactamases, altered drug targets such as conformational changes in PBPs, decreased bacterial permeability, and increased drug efflux. Objectives: The present study aimed to determine the relationship between the results of polymerase chain reaction identification of the Pbp1a, Pbp2b and Pbp2x genes (penicillin-binding proteins) and susceptibilities of β-lactam antibiotics against S. pneumoniae. Materials and Methods: Fifty five isolates of S. pneumoniae were obtained from clinical samples with antimicrobial tests. The susceptibilities of isolates to benzylpenicillin, imipenem, oxacillin, ceftazidime were determined. The resistance genotype was determined by the polymerase chain reaction with primers designed for the PBP genes. Results: The number of S. pneumoniae isolates resistant to benzylpenicillin, imipenem, oxacillin and ceftazidime were 94.5%, 100%, 100%, and 21.8%, respectively. Analysis of mutation in the genes for pbp showed that 85% of isolates had mutations in pbp2x, pbp2b and pbp1a. Susceptibility to benzylpenicillin was decreased once the number of mutated pbp genes in S. pneumonia increased. According to the results of this study, S. pneumoniae isolates showed reduced susceptibility due to accumulation of resistance genes. Conclusions: We suggest that studies should be performed to evaluate changes in Minimum Inhibitory Concentration (MIC) values as well as genetic mutations in order to determine prevalence of S. pneumoniae resistance against antimicrobial agents. PMID:25632328

  2. [Increased susceptibility to non-beta-lactam antimicrobial agents of MRSA isolates: relationship between genotype and antibiotype].

    PubMed

    Bertrand, X; Muller, A; Thouverez, M; Talon, D

    2004-10-01

    The objective of our study was to investigate the relationship between molecular epidemiology and antibiotic susceptibility of MRSA during a four-year period. In this aim, we determined the antibiotype of all MRSA identified during a yearly period of 3 months and typed 50 consecutive non-replicate MRSA isolates of each year. We also recorded antibiotic use. Susceptibility rates to gentamicin, tobramycin and ofloxacin remained stable, respectively, 95, 16 and 4%. In contrast, the proportion of MRSA susceptible to erythromycin progressively increased from 7.0% to 32.5% (P < 0.001). PFGE analysis of genomic DNA from the 200 isolates revealed 15 different clones. We identified two epidemic clones, which contained 150 (clone A) and 28 isolates (clone C), respectively. The proportion of isolates belonging to clone A decreased during the study from 86% to 66%. Conversely, clone C increased from 4% to 22%. The increase of erythromycin-susceptibility within MRSA was caused by the emergence of clone C. Non-epidemic strains were more frequently susceptible to ofloxacin (31.8% vs. 1.1%) and tobramycin (45.4% vs. 16.8%) than epidemic strains. Antimicrobial use had not significantly varied during the study. The proportion of beta-lactams, fluoroquinolones, macrolides and aminosides was 71.8, 13.9, 5.0 and 3.8% of the total antibiotic use, respectively. In our hospital, MRSA isolates became more susceptible to antimicrobial of minor use. The selection pressure exerted by beta-lactams and fluoroquinolones was in favor of the spread of strains resistant to these both major antibiotic classes.

  3. Identification of aminoglycoside and β-lactam resistance genes from within an infant gut functional metagenomic library.

    PubMed

    Fouhy, Fiona; Ogilvie, Lesley A; Jones, Brian V; Ross, R Paul; Ryan, Anthony C; Dempsey, Eugene M; Fitzgerald, Gerald F; Stanton, Catherine; Cotter, Paul D

    2014-01-01

    The infant gut microbiota develops rapidly during the first 2 years of life, acquiring microorganisms from diverse sources. During this time, significant opportunities exist for the infant to acquire antibiotic resistant bacteria, which can become established and constitute the infant gut resistome. With increased antibiotic resistance limiting our ability to treat bacterial infections, investigations into resistance reservoirs are highly pertinent. This study aimed to explore the nascent resistome in antibiotically-naïve infant gut microbiomes, using a combination of metagenomic approaches. Faecal samples from 22 six-month-old infants without previous antibiotic exposure were used to construct a pooled metagenomic library, which was functionally screened for ampicillin and gentamicin resistance. Our library of ∼220Mb contained 0.45 ampicillin resistant hits/Mb and 0.059 gentamicin resistant hits/Mb. PCR-based analysis of fosmid clones and uncloned metagenomic DNA, revealed a diverse and abundant aminoglycoside and β-lactam resistance reservoir within the infant gut, with resistance determinants exhibiting homology to those found in common gut inhabitants, including Escherichia coli, Enterococcus sp., and Clostridium difficile, as well as to genes from cryptic environmental bacteria. Notably, the genes identified differed from those revealed when a sequence-driven PCR-based screen of metagenomic DNA was employed. Carriage of these antibiotic resistance determinants conferred substantial, but varied (2-512x), increases in antibiotic resistance to their bacterial host. These data provide insights into the infant gut resistome, revealing the presence of a varied aminoglycoside and β-lactam resistance reservoir even in the absence of selective pressure, confirming the infant resistome establishes early in life, perhaps even at birth.

  4. Characterization and Prevalence of the Different Mechanisms of Resistance to Beta-Lactam Antibiotics in Clinical Isolates of Escherichia coli

    PubMed Central

    Medeiros, Antone A.; Kent, Ralph L.; O'Brien, Thomas F.

    1974-01-01

    A survey of clinical isolates from a hospital laboratory showed that Escherichia coli could be grouped into three classes of beta-lactam-antibiotic resistance by results of routine susceptibility testing to ampicillin, cephalothin, and carbenicillin. E. coli highly resistant to ampicillin and carbenicillin but not to cephalothin (class I) were found to have one of two levels of R factor-mediated, periplasmic-β-lactamase which resembled RTEM and was located behind a permeability barrier to penicillins but not to cephalosporins. This permeability barrier appeared to act synergistically with the β-lactamase in producing high levels of resistance to penicillins. E. coli highly resistant to ampicillin and cephalothin but not carbenicillin (class II) were found to have a β-lactamase with predominantly cephalosporinase activity which was neither transferable nor releasable by osmotic shock. E. coli moderately resistant to one or to all three of these antibiotics (class III) were found to have low levels of different β-lactamases including a transferable β-lactamase which resembled R1818. Thus, different mechanisms producing resistance to β-lactam antibiotics could be deduced from the patterns of resistance to ampicillin, cephalothin, and carbenicillin found on routine susceptibility testing. E. coli of class I were much more prevalent than the other classes and the proportion of E. coli that were class I increased with duration of patient hospitalization. The incidence of class I E. coli rose only slightly over the past 7 years and that of class II E. coli remained constant despite increased usage of both cephalothin and ampicillin. These observations emphasize that the properties of the apparently limited number of individual resistance mechanisms that exist in a bacterial flora, such as their genetic mobility and linkages and the spectrum of their antibiotic inactivating enzymes and permeability barriers, may govern the effect that usage of an antibiotic has upon

  5. Staphylococcal Phenotypes Induced by Naturally Occurring and Synthetic Membrane-Interactive Polyphenolic β-Lactam Resistance Modifiers

    PubMed Central

    Palacios, Lucia; Rosado, Helena; Micol, Vicente; Rosato, Adriana E.; Bernal, Patricia; Arroyo, Raquel; Grounds, Helen; Anderson, James C.; Stabler, Richard A.; Taylor, Peter W.

    2014-01-01

    Galloyl catechins, in particular (-)-epicatechin gallate (ECg), have the capacity to abrogate β-lactam resistance in methicillin-resistant strains of Staphylococcus aureus (MRSA); they also prevent biofilm formation, reduce the secretion of a large proportion of the exoproteome and induce profound changes to cell morphology. Current evidence suggests that these reversible phenotypic traits result from their intercalation into the bacterial cytoplasmic membrane. We have endeavoured to potentiate the capacity of ECg to modify the MRSA phenotype by stepwise removal of hydroxyl groups from the B-ring pharmacophore and the A:C fused ring system of the naturally occurring molecule. ECg binds rapidly to the membrane, inducing up-regulation of genes responsible for protection against cell wall stress and maintenance of membrane integrity and function. Studies with artificial membranes modelled on the lipid composition of the staphylococcal bilayer indicated that ECg adopts a position deep within the lipid palisade, eliciting major alterations in the thermotropic behaviour of the bilayer. The non-galloylated homolog (-)-epicatechin enhanced ECg-mediated effects by facilitating entry of ECg molecules into the membrane. ECg analogs with unnatural B-ring hydroxylation patterns induced higher levels of gene expression and more profound changes to MRSA membrane fluidity than ECg but adopted a more superficial location within the bilayer. ECg possessed a high affinity for the positively charged staphylococcal membrane and induced changes to the biophysical properties of the bilayer that are likely to account for its capacity to disperse the cell wall biosynthetic machinery responsible for β-lactam resistance. The ability to enhance these properties by chemical modification of ECg raises the possibility that more potent analogs could be developed for clinical evaluation. PMID:24699700

  6. Synergy of β-Lactams with Vancomycin against Methicillin-Resistant Staphylococcus aureus: Correlation of Disk Diffusion and Checkerboard Methods.

    PubMed

    Sy, Cheng Len; Huang, Tsi-Shu; Chen, Chii Shiang; Chen, Yao-Shen; Tsai, Hung-Chin; Wann, Shue-Renn; Wu, Kuan-Sheng; Chen, Jui-Kuang; Lee, Susan Shin-Jung; Liu, Yung-Ching

    2016-03-01

    Modified disk diffusion (MDD) and checkerboard tests were employed to assess the synergy of combinations of vancomycin and β-lactam antibiotics for 59 clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and Mu50 (ATCC 700699). Bacterial inocula equivalent to 0.5 and 2.0 McFarland standard were inoculated on agar plates containing 0, 0.5, 1, and 2 μg/ml of vancomycin. Oxacillin-, cefazolin-, and cefoxitin-impregnated disks were applied to the surface, and the zones of inhibition were measured at 24 h. The CLSI-recommended checkerboard method was used as a reference to detect synergy. The MICs for vancomycin were determined using the Etest method, broth microdilution, and the Vitek 2 automated system. Synergy was observed with the checkerboard method in 51% to 60% of the isolates when vancomycin was combined with any β-lactam. The fractional inhibitory concentration indices were significantly lower in MRSA isolates with higher vancomycin MIC combinations (P < 0.05). The overall agreement between the MDD and checkerboard methods to detect synergy in MRSA isolates with bacterial inocula equivalent to McFarland standard 0.5 were 33.0% and 62.5% for oxacillin, 45.1% and 52.4% for cefazolin, and 43.1% and 52.4% for cefoxitin when combined with 0.5 and 2 μg/ml of vancomycin, respectively. Based on our study, the simple MDD method is not recommended as a replacement for the checkerboard method to detect synergy. However, it may serve as an initial screening method for the detection of potential synergy when it is not feasible to perform other labor-intensive synergy tests. PMID:26677253

  7. Charm Safe-Level beta-Lactam Test for amoxicillin, ampicillin, ceftiofur, cephapirin, and penicillin G in raw commingled milk.

    PubMed

    Salter, R S; Legg, D; Ossanna, N; Boyer, C; Scheemaker, J; Markovsky, R; Saul, S J

    2001-01-01

    The Charm Safe-Level beta-Lactam Test was evaluated by a U.S. Food and Drug Administration (FDA) test protocol administered by the AOAC-Research Institute. The sensitivity and selectivity of the test were evaluated with >800 negative raw commingled and drug-fortified milk samples by the manufacturer and an independent laboratory. Probit analysis by the independent laboratory determined the following 90% positive levels with 95% confidence: amoxicillin, 5.6 ppb; ampicillin, 8.5 ppb; cephapirin, 13.7 ppb; ceftiofur, 46.2 ppb; and penicillin G, 3.6 ppb. These values were within a range of +/- 20% of the manufacturer's data. Selection of negative samples met confidence specifications. Ruggedness parameters were studied and defined, and the stability of frozen milk was verified. There were no interferences from somatic cells (1,000,000 somatic cell count/mL) or bacteria (300,000 colony-forming units/mL), or from 27 other non-beta-lactam animal drugs. Test performance with raw milk samples containing incurred penicillin, ampicillin, and amoxicillin was consistent with the dose responses determined with fortified milk samples. Incurred cephalosporin in raw milk samples was detected at lower levels than was cephalosporin in fortified milk samples, presumably because of the presence of metabolite, as verified by other test methods. Quality control data support consistency in manufacture between batches and the stability of refrigerated test reagents for up to 1 year. Successful fulfillment of these criteria led to FDA certification of the test when used with a reader in U.S. milk testing programs.

  8. Efficacy of β-Lactam-plus-Macrolide Combination Therapy in a Mouse Model of Lethal Pneumococcal Pneumonia.

    PubMed

    Yoshioka, Daisuke; Kajiwara, Chiaki; Ishii, Yoshikazu; Umeki, Kenji; Hiramatsu, Kazufumi; Kadota, Jun-Ichi; Tateda, Kazuhiro

    2016-10-01

    Community-acquired pneumonia is a common disease with considerable morbidity and mortality, for which Streptococcus pneumoniae is accepted as a leading cause. Although β-lactam-plus-macrolide combination therapy for this disease is recommended in several guidelines, the clinical efficacy of this strategy against pneumococcal pneumonia remains controversial. In this study, we examined the effects of β-lactam-plus-macrolide combination therapy on lethal mouse pneumococcal pneumonia and explored the mechanisms of action in vitro and in vivo We investigated survival, lung bacterial burden, and cellular host responses in bronchoalveolar lavage fluids obtained from mice infected with pneumonia and treated with ceftriaxone, azithromycin, or both in combination. Although in vitro synergy was not observed, significant survival benefits were demonstrated with combination treatment. Lung neutrophil influx was significantly lower in the ceftriaxone-plus-azithromycin-treated group than in the ceftriaxone-treated group, whereas no differences in the lung bacterial burden were observed on day 3 between the ceftriaxone-plus-azithromycin-treated group and the ceftriaxone-treated group. Notably, the analysis of cell surface markers in the ceftriaxone-plus-azithromycin combination group exhibited upregulation of presumed immune checkpoint ligand CD86 and major histocompatibility complex class II in neutrophils and CD11b-positive CD11c-positive (CD11b(+) CD11c(+)) macrophages and dendritic cells, as well as downregulation of immune checkpoint receptors cytotoxic-T lymphocyte-associated antigen 4 and programmed death 1 in T helper and T regulatory cells. Our data demonstrate that the survival benefits of ceftriaxone-plus-azithromycin therapy occur through modulation of immune checkpoints in mouse pneumococcal pneumonia. In addition, immune checkpoint molecules may be a novel target class for future macrolide research. PMID:27480866

  9. Interaction of β-lactam carbenes with 3,6-diphenyltetrazines: a five-step cascade reaction for the direct construction of indeno[2,1-b]pyrrol-2-ones.

    PubMed

    Xing, Juan; Wang, Xiao-Rong; Yan, Cai-Xia; Cheng, Ying

    2011-06-01

    A study of the nucleophilic addition of β-lactam carbenes to 3,6-diphenyltetrazines is reported. Instead of the formation of pyrazole derivatives like most reactions between nucleophilic or ambiphilic carbenes and 3,6-disubstituted tetrazines, β-lactam carbenes reacted with 3,6-diphenyltetrazines to produce indeno[2,1-b]pyrrol-2-ones in good yields. The reaction proceeds most probably through a five-step cascade process. This work has not only provided a one-pot operation for the efficient construction of mutisubstituted indeno[2,1-b]pyrrol-2-ones but also revealed the nucleophilicity of β-lactam carbenes.

  10. Role of Pseudomonas aeruginosa low-molecular-mass penicillin-binding proteins in AmpC expression, β-lactam resistance, and peptidoglycan structure.

    PubMed

    Ropy, Alaa; Cabot, Gabriel; Sánchez-Diener, Irina; Aguilera, Cristian; Moya, Bartolome; Ayala, Juan A; Oliver, Antonio

    2015-07-01

    This study aimed to characterize the role of Pseudomonas aeruginosa low-molecular-mass penicillin-binding proteins (LMM PBPs), namely, PBP4 (DacB), PBP5 (DacC), and PBP7 (PbpG), in peptidoglycan composition, β-lactam resistance, and ampC regulation. For this purpose, we constructed all single and multiple mutants of dacB, dacC, pbpG, and ampC from the wild-type P. aeruginosa PAO1 strain. Peptidoglycan composition was determined by high-performance liquid chromatography (HPLC), ampC expression by reverse transcription-PCR (RT-PCR), PBP patterns by a Bocillin FL-binding test, and antimicrobial susceptibility by MIC testing for a panel of β-lactams. Microscopy and growth rate analyses revealed no apparent major morphological changes for any of the mutants compared to the wild-type PAO1 strain. Of the single mutants, only dacC mutation led to significantly increased pentapeptide levels, showing that PBP5 is the major dd-carboxypeptidase in P. aeruginosa. Moreover, our results indicate that PBP4 and PBP7 play a significant role as dd-carboxypeptidase only if PBP5 is absent, and their dd-endopeptidase activity is also inferred. As expected, the inactivation of PBP4 led to a significant increase in ampC expression (around 50-fold), but, remarkably, the sequential inactivation of the three LMM PBPs produced a much greater increase (1,000-fold), which correlated with peptidoglycan pentapeptide levels. Finally, the β-lactam susceptibility profiles of the LMM PBP mutants correlated well with the ampC expression data. However, the inactivation of ampC in these mutants also evidenced a role of LMM PBPs, especially PBP5, in intrinsic β-lactam resistance. In summary, in addition to assessing the effect of P. aeruginosa LMM PBPs on peptidoglycan structure for the first time, we obtained results that represent a step forward in understanding the impact of these PBPs on β-lactam resistance, apparently driven by the interplay between their roles in AmpC induction, β-lactam

  11. Distinct molecular structures and hydrogen bond patterns of α,α-diethyl-substituted cyclic imide, lactam, and acetamide derivatives in the crystalline phase

    NASA Astrophysics Data System (ADS)

    Krivoshein, Arcadius V.; Ordonez, Carlos; Khrustalev, Victor N.; Timofeeva, Tatiana V.

    2016-10-01

    α,α-Dialkyl- and α-alkyl-α-aryl-substituted cyclic imides, lactams, and acetamides show promising anticonvulsant, anxiolytic, and anesthetic activities. While a number of crystal structures of various α-substituted cyclic imides, lactams, and acetamides were reported, no in-depth comparison of crystal structures and solid-state properties of structurally matched compounds have been carried out so far. In this paper, we report molecular structure and intermolecular interactions of three α,α-diethyl-substituted compounds - 3,3-diethylpyrrolidine-2,5-dione, 3,3-diethylpyrrolidin-2-one, and 2,2-diethylacetamide - in the crystalline phase, as studied using single-crystal X-ray diffraction and IR spectroscopy. We found considerable differences in the patterns of H-bonding and packing of the molecules in crystals. These differences correlate with the compounds' melting points and are of significance to physical pharmacy and formulation development of neuroactive drugs.

  12. Structures, semisyntheses, and absolute configurations of the antiplasmodial α-substituted β-lactam monamphilectines B and C from the sponge Svenzea flava

    PubMed Central

    Avilés, Edward; Prudhomme, Jacques; Le Roch, Karine G.

    2014-01-01

    Bioassay-guided fractionation of the Caribbean sponge Svenzea flava collected near Mona Island, off the west coast of Puerto Rico, led to the isolation of two isocyanide amphilectane-type diterpenes named monamphilectines B and C (2 and 3). Attached to the backbone of each of these compounds is the first α-substituted monocyclic β-lactam ring to be isolated from a marine organism. The molecular structures of 2 and 3 were established by spectroscopic methods and then confirmed unequivocally by chemical correlation and comparison of physical and chemical data with the natural products. The new β-lactams were successfully synthesized in one step, starting from the known diisocyanide 4, via parallel Ugi four-center three-component reactions (U-4C-3CR) that also established their absolute stereostructures. Interestingly, compounds 2 and 3 exhibited activities in the low nanomolar range against the human malaria parasite Plasmodium falciparum. PMID:26494928

  13. Stereocontrolled Annulations of Indolo[2,3-a]quinolizidine-Derived Lactams with a Silylated Nazarov Reagent: Access to Allo and Epiallo Yohimbine-Type Derivatives.

    PubMed

    Arioli, Federica; Pérez, Maria; Are, Celeste; Estarellas, Carolina; Luque, F Javier; Bosch, Joan; Amat, Mercedes

    2015-09-14

    The facial selectivity of double Michael addition reactions of the silylated Nazarov reagent 4 to unsaturated indolo[2,3-a]quinolizidine lactams 3 has been studied. Pentacyclic 3-H/15-H trans adducts 5 are generated from Nind -unsubstituted lactams, but the corresponding cis isomers 6 are formed when the indole nitrogen has a tert-butyloxycarbonyl (Boc) substituent. This reversal in the facial selectivity of the annulation has been rationalized by means of theoretical calculations, which indicate that the initial nucleophilic attack under stereoelectronic control is hampered by the presence of the bulky Boc group. The synthetic usefulness of the pentacyclic Nazarov-derived adducts is demonstrated by their conversion into allo and epiallo yohimbine-type targets. PMID:26332232

  14. Salicylate decreases production of AmpC type beta-lactamases and increases susceptibility to beta-lactams in a Morganella morganii clinical isolate.

    PubMed

    Tavío, María M; Perilli, Mariagrazia; Vila, Jordi; Becerro, Pino; Casañas, Lucía; Amicosante, Gianfranco; Jiménez de Anta, María Teresa

    2004-09-01

    The effect of salicylate, a marRAB inducer, on the resistance to beta-lactams was characterized in an AmpC beta-lactamase hyperproducer Morganella morganii clinical isolate (the M1 strain). Results were compared with those of the effect of salicylate in a wild-type M. morganii strain. Salicylate induced a decreased susceptibility to nalidixic acid, norfloxacin and tetracycline and simultaneously increased the susceptibility to beta-lactams apparently due to the repression of AmpC beta-lactamase synthesis in the M1 strain. Likewise, salicylate only repressed 46 kDa outer membrane protein expression in the wild-type strain, since the clinical isolate M1 did not express it.

  15. Molecular mechanisms involved in interleukin 1-beta (IL-1β)-induced memory impairment. Modulation by alpha-melanocyte-stimulating hormone (α-MSH).

    PubMed

    Gonzalez, P; Machado, I; Vilcaes, A; Caruso, C; Roth, G A; Schiöth, H; Lasaga, M; Scimonelli, T

    2013-11-01

    Pro-inflammatory cytokines can affect cognitive processes such as learning and memory. Particularly, interleukin-1β (IL-1β) influences the consolidation of hippocampus-dependent memories. We previously reported that administration of IL-1β in dorsal hippocampus impaired contextual fear memory consolidation. Different mechanisms have been implicated in the action of IL-1β on long-term potentiation (LTP), but the processes by which this inhibition occurs in vivo remain to be elucidated. We herein report that intrahippocampal injection of IL-1β induced a significant increase in p38 phosphorylation after contextual fear conditioning. Also, treatment with SB203580, an inhibitor of p38, reversed impairment induced by IL-1β on conditioned fear behavior, indicating that this MAPK would be involved in the effect of the cytokine. We also showed that IL-1β administration produced a decrease in glutamate release from dorsal hippocampus synaptosomes and that treatment with SB203580 partially reversed this effect. Our results indicated that IL-1β-induced impairment in memory consolidation could be mediated by a decrease in glutamate release. This hypothesis is sustained by the fact that treatment with d-cycloserine (DCS), a partial agonist of the NMDA receptor, reversed the effect of IL-1β on contextual fear memory. Furthermore, we demonstrated that IL-1β produced a temporal delay in ERK phosphorylation and that DCS administration reversed this effect. We also observed that intrahippocampal injection of IL-1β decreased BDNF expression after contextual fear conditioning. We previously demonstrated that α-MSH reversed the detrimental effect of IL-1β on memory consolidation. The present results demonstrate that α-MSH administration did not modify the decrease in glutamate release induced by IL-1β. However, intrahippocampal injection of α-MSH prevented the effect on ERK phosphorylation and BDNF expression induced by IL-1β after contextual fear conditioning. Therefore, in the present study we determine possible molecular mechanisms involved in the impairment induced by IL-1β on fear memory consolidation. We also established how this effect could be modulated by α-MSH.

  16. Linker modification reduced the renal uptake of technetium-99m-labeled Arg-Ala-Asp-conjugated alpha-melanocyte stimulating hormone peptide.

    PubMed

    Yang, Jianquan; Flook, Adam M; Feng, Changjian; Miao, Yubin

    2014-01-01

    The purpose of this study was to examine the biodistribution of (99m)Tc-RAD-Arg-(Arg(11))CCMSH in B16/F1 melanoma-bearing C57 mice to determine whether the replacement of the Lys linker with an Arg linker could decrease the renal uptake of (99m)Tc-RAD-Arg-(Arg(11))CCMSH. (99m)Tc-RAD-Arg-(Arg(11))CCMSH exhibited rapid and high tumor uptake (17.98±4.96% ID/g at 2h post-injection) in B16/F1 melanoma-bearing C57 mice. As compared to (99m)Tc-RAD-Lys-(Arg(11))CCMSH, the replacement of the Lys linker with an Arg linker dramatically decreased the renal uptake of (99m)Tc-RAD-Arg-(Arg(11))CCMSH by 68%, 62%, 73% and 64% at 0.5, 2, 4 and 24h post-injection, respectively. Flank B16/F1 melanoma lesions were clearly imaged at 2h post-injection using (99m)Tc-RAD-Arg-(Arg(11))CCMSH as an imaging probe.

  17. Therapeutic Efficacy of a {sup 188}Re-Labeled {alpha}-Melanocyte-Stimulating Hormone Peptide Analog in Murine and Human Melanoma-Bearing Mouse Models

    SciTech Connect

    Miao, Yubin; Owen, Nellie K.; Fisher, Darrell R.; Hoffman, Timothy J.; Quinn, Thomas P.

    2005-01-01

    The purpose of this study was to examine the therapeutic efficacy of {sup 188}Re-(Arg{sup 11})CCMSH in the B16/F1 murine melanoma and TXM13 human melanoma bearing mouse models. Method: (Arg11)CCMSH was synthesized and labeled with {sup 188}Re to form {sup 188}Re-(Agr{sup 11})CCMSH. B16/F1 melanoma tumor bearing mice were administrated with 200 Ci, 600 Ci and 2x400 Ci of {sup 188}Re-(Arg{sup 11})CCMSH via the tail vein, respectively. TXM13 melanoma tumor hearing mice were separately injected with 600 Ci, 2x400 Ci and 1000 Ci of 100Re-(Arg{sup 11})CCMSH through the tail vein. Two groups of 10 mice bearing either B16/F1 or TXM13 tumors were injected with saline as untreated controls. Results: In contrast to the untreated control group, {sup 188}Re(Arg11)CCMSH yielded rapid and lasting therapeutic effects in the treatment groups with either B16/F1 or TXM13 tumors. The tumor growth rate was reduced and the survival rate was prolonged in the treatment groups. Treatment with 2x400 Ci of {sup 188}Re-Arg{sup 11}CCMSH significantly extended the mean life of B16/F1 tumor mice (p<0.05), while the mean life of TXm13 tumor mice was significantly prolonged after treatment with 600 Ci and 1000 Ci doses of {sup 188}Re-(Arg{sup 11})CCMSH (p<0.05 High-dose {sup 188}Re-(Arg{sup 11}))CCMSH produced no observed normal-tissue toxicity. Conclusions: The therapy study results revealed that {sup 188}Re-Arg11 CCMSH yielded significant therapeutic effects in both B16/F1 murine melanoma and TXM13 human melanoma bearing mouse models. {sup 188}Re-(Arg{sup 11})CCMSH appears to be a promising radiolabeled peptide for targeted radionuclide therapy of melanoma.

  18. Validation of a rapid lateral flow test for the simultaneous determination of β-lactam drugs and flunixin in raw milk.

    PubMed

    Douglas, David; Banaszewski, Katie; Juskelis, Rima; Al-Taher, Fadwa; Chen, Yang; Cappozzo, Jack; McRobbie, Lindsay; Salter, Robert S

    2012-07-01

    β-Lactam antibiotics are the most commonly used drugs on dairy farms. β-Lactam residues in milk are kept out of the human milk supply with good agricultural practices and mandatory truck screening performed by the dairy industry under Appendix N of the Pasteurized Milk Ordinance. Flunixin, a nonsteroidal and anti-inflammatory drug, appears in dairy cattle tissue residues with a frequency similar to the occurrence of penicillin G. This creates concern that flunixin residues could be in milk and would go undetected under current milk screening programs. A single test that combines mandatory β-lactam screening with voluntary flunixin screening is an economical approach for monitoring and controlling for potential flunixin or 5-hydroxyflunixin, the primary flunixin metabolite marker in milk. The objective of this study was to validate a β-lactam and flunixin rapid lateral flow test (LFT) and compare the results obtained with a liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of flunixin and 5-hydroxyflunixin in raw milk with a limit of detection of , 1 ppb, equivalent to 1 ng/ml. Using the LFT, three combined manufactured lots of test strips detected penicillin G at 2.0 ppb, ampicillin at 6.8 ppb, amoxicillin at 5.9 ppb, cephapirin at 13.4 ppb, ceftiofur (total metabolites) at 63 ppb, and 5-hydroxyflunixin at 1.9 ppb at least 90% of the time with 95% confidence. The LFT also detected incurred flunixin milk samples that were analyzed with the LC-MS/MS and diluted to tolerance in raw milk. The detection levels for the LFT are lower than the U.S. safe levels or tolerances and qualify the test to be used in compliance with U.S. milk screening programs.

  19. The Beta Lactam Antibiotics as an Empirical Therapy in a Developing Country: An Update on Their Current Status and Recommendations to Counter the Resistance against Them

    PubMed Central

    Thakuria, Bhaskar; Lahon, Kingshuk

    2013-01-01

    In a developing country like India, where the patients have to bear the cost of their healthcare, the microbiological culture and the sensitivity testing of each and every infection is not feasible. Moreover, there are lacunae in the data storage, management and the sharing of knowledge with respect to the microorganisms which are prevalent in the local geographical area and with respect to the antibiotics which are effective against them. Thus, an empirical therapy for treating infections is imperative in such a setting. The beta lactam antibiotics have been widely used for the empirical treatment of infections since the the discovery of penicillin. Many generations of beta lactams have been launched with, the claims of a higher sensitivity and less resistance, but their sensitivity has drastically decreased over time. Thus, the preference for beta lactams, especially the cephalosporins, as an empirical therapy, among the prescribers was justified initially, but the current sensitivity patterns do not support their empirical use in hospital and community acquired infections. There is a need for increasing the awareness and the attitudinal change among the prescribers, screening of the antibiotic prescriptions, the strict implementation of antibiotic policies in hospital settings, restricting the hospital supplies and avoiding the prescriptions of beta lactams, a regular census of the local sensitivity patterns to formulate and update the antibiotic policies, upgradation of the laboratory facilities for a better and faster detection of the isolates, proper collection, analyses and sharing of the data and the encouragement of the research and development of newer antibiotics with novel mechanisms of action. PMID:23905143

  20. An N-Heterocyclic Carbene-Catalyzed Oxidative γ-Aminoalkylation of Saturated Carboxylic Acids through in Situ Activation Strategy: Access to δ-Lactam.

    PubMed

    Que, Yonglei; Xie, Yuanwei; Li, Tuanjie; Yu, Chenxia; Tu, Shujiang; Yao, Changsheng

    2015-12-18

    An N-Heterocyclic Carbene (NHC)-catalyzed oxidative formal [4 + 2] annulation of acylhydrazones with saturated carboxylic acids bearing γ-H to assemble δ-lactams featuring a chiral carbon stereogenic center was developed through an in situ activation strategy. The ready availability of the starting materials, excellent enantioselectivity, facile assembly, high yields, and potential biological significance of the final products make this protocol an attractive alternative for the construction of the pyridinone scaffold. PMID:26646554

  1. Validation of a rapid lateral flow test for the simultaneous determination of β-lactam drugs and flunixin in raw milk.

    PubMed

    Douglas, David; Banaszewski, Katie; Juskelis, Rima; Al-Taher, Fadwa; Chen, Yang; Cappozzo, Jack; McRobbie, Lindsay; Salter, Robert S

    2012-07-01

    β-Lactam antibiotics are the most commonly used drugs on dairy farms. β-Lactam residues in milk are kept out of the human milk supply with good agricultural practices and mandatory truck screening performed by the dairy industry under Appendix N of the Pasteurized Milk Ordinance. Flunixin, a nonsteroidal and anti-inflammatory drug, appears in dairy cattle tissue residues with a frequency similar to the occurrence of penicillin G. This creates concern that flunixin residues could be in milk and would go undetected under current milk screening programs. A single test that combines mandatory β-lactam screening with voluntary flunixin screening is an economical approach for monitoring and controlling for potential flunixin or 5-hydroxyflunixin, the primary flunixin metabolite marker in milk. The objective of this study was to validate a β-lactam and flunixin rapid lateral flow test (LFT) and compare the results obtained with a liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of flunixin and 5-hydroxyflunixin in raw milk with a limit of detection of , 1 ppb, equivalent to 1 ng/ml. Using the LFT, three combined manufactured lots of test strips detected penicillin G at 2.0 ppb, ampicillin at 6.8 ppb, amoxicillin at 5.9 ppb, cephapirin at 13.4 ppb, ceftiofur (total metabolites) at 63 ppb, and 5-hydroxyflunixin at 1.9 ppb at least 90% of the time with 95% confidence. The LFT also detected incurred flunixin milk samples that were analyzed with the LC-MS/MS and diluted to tolerance in raw milk. The detection levels for the LFT are lower than the U.S. safe levels or tolerances and qualify the test to be used in compliance with U.S. milk screening programs. PMID:22980011

  2. Trace determination of beta-lactam antibiotics in surface water and urban wastewater using liquid chromatography combined with electrospray tandem mass spectrometry.

    PubMed

    Cha, J M; Yang, S; Carlson, K H

    2006-05-19

    A sensitive and reliable method using liquid chromatography-electrospray tandem mass spectrometry has been developed and validated for the trace determination of beta-lactam antibiotics in natural and wastewater matrices. Water samples were enriched by solid-phase extraction. The analytes included amoxicillin (AMOX), ampicillin (AMP), oxacillin (OXA), cloxacillin (CLOX) and cephapirin (CEP). Average recoveries of beta-lactams (BLs) in fortified samples were generally above 75% (except amoxicillin) with the standard deviations lower than 10% in water matrices. Amoxicillin was not quantified due to poor recovery (less than 40%) in the investigated water matrices. Matrix effects were found to be minimal when measuring these compounds in water matrices. The accuracy, within- and between-run precision of the assay fell within acceptable ranges of 15% absolute. The method detection limit (MDL) was estimated to range between 8 and 10 ng/L in surface water, 13 and 18 ng/L in the influent and 8 and 15 ng/L in the effluent from a wastewater treatment plant. A large number of actual water samples were analyzed using this method in order to evaluate the occurrence of the beta-lactams in a river and a wastewater treatment plant in northern Colorado. Most of the samples were negative for all analytes. These compounds were found at 15-17 ng/L in the three influent samples and at 9-11 ng/L in three surface water samples out of a total of 200 samples. This indicates that contamination by beta-lactam antibiotics is of minor importance to the small mixed-watershed. PMID:16595135

  3. Tin Powder-Promoted One-Pot Construction of α-Methylene-γ-lactams and Spirolactams from Aldehydes or Ketones, Acylhydrazines, and 2-(Bromomethyl)acrylate.

    PubMed

    Xu, Yanli; Huang, Danfeng; Wang, Ke-Hu; Ma, Junyan; Su, Yingpeng; Fu, Ying; Hu, Yulai

    2015-12-18

    A concise and efficient method for the synthesis of α-methylene-γ-lactams is developed from multicomponent one-pot reactions of aldehydes or ketones, hydrazides, and ethyl 2-(bromomethyl)acrylate promoted by tin powder. The reaction proceeds smoothly under mild reaction conditions without using any catalyst to give the corresponding products in high yields. α-Methylene-γ-spirolactams can also be prepared from cyclic ketones. PMID:26577682

  4. Highly stable atropisomers by electrophilic amination of a chiral γ-lactam within the synthesis of an elusive conformationally restricted analogue of α-methylhomoserine.

    PubMed

    Amabili, Paolo; Amici, Adolfo; Civitavecchia, Annafelicia; Maggiore, Beatrice; Orena, Mario; Rinaldi, Samuele; Tolomelli, Alessandra

    2016-02-01

    Starting from chiral-protected 4-hydroxymethyl pyrrolidin-2-ones, the otherwise elusive 3,4-trans-3,3,4-trisubstituted isosteres of α-methyl homoserine, tethered on a γ-lactam ring, were prepared exploiting stereoselective electrophilic aminations. These reactions led to the isolation and characterization of a novel type of atropisomers, exceedingly stable at room temperature, that were directly converted to the desired products by a novel non-reductive N-N bond cleavage reaction.

  5. Diastereoselective Synthesis of syn-β-Lactams Using Rh-Catalyzed Reductive Mannich-Type Reaction of α,β-Unsaturated Esters.

    PubMed

    Isoda, Motoyuki; Sato, Kazuyuki; Funakoshi, Masato; Omura, Keiko; Tarui, Atsushi; Omote, Masaaki; Ando, Akira

    2015-08-21

    The combination of Et2Zn and RhCl(PPh3)3 led to the facile generation of a rhodium-hydride complex (Rh-H) that catalyzed the 1,4-reduction of α,β-unsaturated esters. The resulting rhodium enolate performed as a Reformatsky-type reagent and reacted with various imines to give syn-β-lactams in good to excellent yields with high diastereoselectivity. PMID:26203668

  6. Multicenter evaluation of the clinical outcomes of daptomycin with and without concomitant β-lactams in patients with Staphylococcus aureus bacteremia and mild to moderate renal impairment.

    PubMed

    Moise, Pamela A; Amodio-Groton, Maria; Rashid, Mohamad; Lamp, Kenneth C; Hoffman-Roberts, Holly L; Sakoulas, George; Yoon, Min J; Schweitzer, Suzanne; Rastogi, Anjay

    2013-03-01

    Patients with underlying renal disease may be vulnerable to vancomycin-mediated nephrotoxicity and Staphylococcus aureus bacteremia treatment failure. In light of recent data demonstrating the successful use of β-lactam plus daptomycin in very difficult cases of S. aureus bacteremia, we examined safety and clinical outcomes for patients who received daptomycin with or without concomitant β-lactams. We identified 106 patients who received daptomycin for S. aureus bacteremia, had mild or moderate renal insufficiency according to FDA criteria, and enrolled in the Cubicin Outcomes Registry and Experience (CORE), a multicenter registry, from 2005 to 2009. Daptomycin treatment success was 81%. Overall treatment efficacy was slightly enhanced with the addition of a β-lactam (87% versus 78%; P = 0.336), but this trend was most pronounced for bacteremia associated with endocarditis or bone/joint infection or bacteremia from an unknown source (90% versus 57%; P = 0.061). Factors associated with reduced daptomycin efficacy (by logistic regression) were an unknown source of bacteremia (odds ratio [OR] = 7.59; 95% confidence interval [CI] = 1.55 to 37.2), moderate renal impairment (OR = 9.11; 95% CI = 1.46 to 56.8), and prior vancomycin failure (OR = 11.2; 95% CI = 1.95 to 64.5). Two patients experienced an increase in creatine phosphokinase (CPK) that resolved after stopping daptomycin. No patients developed worsening renal insufficiency related to daptomycin. In conclusion, daptomycin appeared to be effective and well tolerated in patients with S. aureus bacteremia and mild to moderate renal insufficiency. Daptomycin treatment efficacy might be enhanced with β-lactam combination therapy in primary endovascular and bone/joint infections. Additional studies will be necessary to confirm these findings. PMID:23254428

  7. In-silico modeling of a novel OXA-51 from β-lactam-resistant Acinetobacter baumannii and its interaction with various antibiotics.

    PubMed

    Tiwari, Vishvanath; Nagpal, Isha; Subbarao, Naidu; Moganty, Rajeswari R

    2012-07-01

    Acinetobacter baumannii, one of the major Gram negative bacteria, causes nosocomial infections such as pneumonia, urinary tract infection, meningitis, etc. β-lactam-based antibiotics like penicillin are used conventionally to treat infections of A. baumannii; however, they are becoming progressively less effective as the bacterium produces diverse types of β-lactamases to inactivate the antibiotics. We have recently identified a novel β-lactamase, OXA-51 from clinical strains of A. baumannii from our hospital. In the present study, we generated the structure of OXA-51 using MODELLER9v7 and studied the interaction of OXA-51 with a number of β-lactams (penicillin, oxacillin, ceftazidime, aztreonam and imipenem) using two independent programs: GLIDE and GOLD. Based on the results of different binding parameters and number of hydrogen bonds, interaction of OXA-51 was found to be maximum with ceftazidime and lowest with imipenem. Further, molecular dynamics simulation results also support this fact. The lowest binding affinity of imipenem to OXA-51 indicates clearly that it is not efficiently cleaved by OXA-51, thus explaining its high potency against resistant A. baumannii. This finding is supported by experimental results from minimum inhibitory concentration analysis and transmission electron microscopy. It can be concluded that carbapenems (imipenem) are presently effective β-lactam antibiotics against resistant strains of A. baumannii harbouring OXA-51. The results presented here could be useful in designing more effective derivatives of carbapenem.

  8. The beta-lactam antibiotic ceftriaxone inhibits physical dependence and abstinence-induced withdrawal from cocaine, amphetamine, methamphetamine, and clorazepate in planarians.

    PubMed

    Rawls, Scott M; Cavallo, Federica; Capasso, Anna; Ding, Zhe; Raffa, Robert B

    2008-04-28

    Ceftriaxone (a beta-lactam antibiotic) has recently been identified as having the rare ability to increase the expression and functional activity of the glutamate transporter subtype 1 (GLT-1) in rat spinal cord cultures. GLT-1 has been implicated in diverse neurological disorders and in opioid dependence and withdrawal. It has been speculated that it might also be involved in the physical dependence and withdrawal of other abused drugs, but demonstration of this property can be difficult in mammalian models. Here, we demonstrate for the first time using a planarian model that ceftriaxone attenuates both the development of physical dependence and abstinence-induced withdrawal from cocaine, amphetamine, methamphetamine, and a benzodiazepine (clorazepate) in a concentration-related manner. These results suggest that physical dependence and withdrawal from several drugs involve a common - beta-lactam-sensitive - mechanism in planarians. If these findings can be shown to extend to mammals, beta-lactam antibiotics might represent a novel pharmacotherapy or adjunct approach for treating drug abuse or serve as a template for drug discovery efforts aimed at treating drug abuse, recovery from drug abuse, or ameliorating the withdrawal from chronic use of therapeutic medications.

  9. Structural studies on 4,5-disubstituted 2-aminoimidazole-based biofilm modulators that suppress bacterial resistance to β-lactams.

    PubMed

    Su, Zhaoming; Yeagley, Andrew A; Su, Rui; Peng, Lingling; Melander, Christian

    2012-11-01

    A library of 4,5-disubstituted 2-aminoimidazole triazole amide (2-AITA) conjugates has been successfully assembled. Upon biological screening, this class of small molecules was discovered as enhanced biofilm regulators through non-microbicidal mechanisms against methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Acinetobacter baumannii (MDRAB), with active concentrations in the low micromolar range. The library was also subjected to synergism and resensitization studies with β-lactam antibiotics against MRSA. Lead compounds were identified that suppress the antibiotic resistance of MRSA by working synergistically with oxacillin, a β-lactam antibiotic resistant to penicillinase. A further structure-activity relationship (SAR) study on the parent 2-AITA compound delivered a 2-aminoimidazole diamide (2-AIDA) conjugate with significantly increased synergistic activity with oxacillin against MRSA, decreasing the MIC value of the β-lactam antibiotic by 64-fold. Increased anti-biofilm activity did not necessarily lead to increased suppression of antibiotic resistance, which indicates that biofilm inhibition and resensitization are most likely occurring via distinct mechanisms. PMID:23011973

  10. The selection of suitable columns for a reversed-phase liquid chromatographic separation of beta-lactam antibiotics and related substances via chromatographic column parameters.

    PubMed

    Zhang, Wei-qing; Hu, Qiu-xin; Zhang, Xia; Li, Ya-ping; Wang, Ming-juan; Hu, Chang-qin

    2014-01-01

    The selection of RP-LC columns suitable for a particular analysis in official compendia is difficult as only a general description of the stationary phase in the description of a LC method is given. General methods to characterize RP-LC columns often assume that each of the column parameters is equally important. This can cause the user to select columns inappropriate for particular analyses. This paper focuses on the relationship between the critical peak pairs and the column parameters (H, S, A, B, and C) in the Snyder/Dolan column characterization methodology to find the key parameters influencing real separations. Some varieties of β-lactam antibiotics and their related compounds were used as test compounds. We found column parameter A to be the most important factor affecting their separation. Parameters B and C also played an important role in some separation processes. This indicated that the hydrogen bonding of column and solute can directly affect the separation of β-lactam antibiotics. Choosing columns for which column parameter A is near 0.1 can facilitate the ideal separations of impurities from β-lactam antibiotics. The most suitable column for any common pharmaceutical analysis could be selected easily if the key column parameters would be given in the description of the chromatographic method. For these reasons, key column parameters should be listed in the monographs of official compendia.

  11. Recognition and binding of β-lactam antibiotics to bovine serum albumin by frontal affinity chromatography in combination with spectroscopy and molecular docking.

    PubMed

    Li, Qian; Zhang, Tianlong; Bian, Liujiao

    2016-03-01

    Serum albumins are the most abundant carrier proteins in blood plasma and participate in the binding and transportation of various exogenous and endogenous compounds in the body. This work was designed to investigate the recognition and binding of three typical β-lactam antibiotics including penicillin G (Pen G), penicillin V (Pen V) and cefalexin (Cef) with bovine serum albumin (BSA) by frontal affinity chromatography in combination with UV-vis absorption spectra, fluorescence emission spectra, binding site marker competitive experiment and molecular docking under simulated physiological conditions. The results showed that a BSA only bound with one antibiotic molecule in the binding process, and the binding constants for Pen G-BSA, Pen V-BSA and Cef-BSA complexes were 4.22×10(1), 4.86×10(2) and 3.32×10(3) (L/mol), respectively. All the three β-lactam antibiotics were mainly inserted into the subdomain IIA (binding site 1) of BSA by hydrogen bonds and Van der Waals forces. The binding capacity between the antibiotics and BSA was closely related to the functional groups and flexibility of side chains in antibiotics. This study provided an important insight into the molecular recognition and binding interaction of BSA with β-lactam antibiotics, which may be a useful guideline for the innovative clinical medications and new antibiotic designs with effective pharmacological properties. PMID:26882128

  12. Label-Free Photonic Crystal-Based β-Lactamase Biosensor for β-Lactam Antibiotic and β-Lactamase Inhibitor.

    PubMed

    Xiao, Fubing; Li, Guoguo; Wu, Yan; Chen, Qianshan; Wu, Zhaoyang; Yu, Ruqin

    2016-09-20

    A simple, label-free, and visual photonic crystal-based β-lactamase biosensor was developed for β-lactam antibiotic and β-lactamase inhibitor in which the penicillinase (a β-lactamase) was immobilized on the pH-sensitive colloidal crystal hydrogel (CCH) film to form penicillinase colloidal crystal hydrogel (PCCH) biosensing film. The hydrolysis of penicillin G (a β-lactam antibiotic) can be catalyzed by penicillinase to produce penicilloic acid, leading to a pH decrease in the microenvironment of PCCH film, which causes the shrink of pH-sensitive CCH film and triggers a blue-shift of the diffraction wavelength. Upon the addition of β-lactamase inhibitor, the hydrolysis reaction is suppressed and no clear blue-shift is observed. The concentrations of β-lactam antibiotic and β-lactamase inhibitor can be sensitively evaluated by measuring the diffraction shifts. The minimum detectable concentrations for penicillin G and clavulanate potassium (a β-lactamase inhibitor) can reach 1 and 0.1 μM, respectively. Furthermore, the proposed method is highly reversible and selective, and it allows determination of penicillin G in fish pond water samples. PMID:27552182

  13. Daptomycin-β-Lactam Combinations in a Rabbit Model of Daptomycin-Nonsusceptible Methicillin-Resistant Staphylococcus aureus Endocarditis.

    PubMed

    Chambers, Henry F; Basuino, Li; Hamilton, Stephanie M; Choo, Eun Ju; Moise, Pamela

    2016-07-01

    Beta-lactams enhance the in vitro activity of daptomycin against methicillin-resistant strains of Staphylococcus aureus Experiments were performed in a rabbit model of aortic valve endocarditis caused by methicillin-resistant daptomycin-nonsusceptible S. aureus strain CB5054 to determine if a cephalosporin, ceftriaxone, administered as a once-daily dose of 100 mg/kg of body weight, or a carbapenem, ertapenem, administered as a once-daily dose of 40 mg/kg, improved the efficacy of daptomycin, administered as a once-daily dose of 12 mg/kg. Daptomycin was ineffective alone in reducing organism densities compared to untreated controls in vegetations and spleen, but densities were 1.4 log10 CFU/g lower in kidney. The combination of daptomycin plus ceftriaxone or daptomycin plus ertapenem reduced bacterial densities in all tissues compared to single agents, with 0.6 to 1.0 log10 CFU/g fewer organisms in vegetations, 1.5 to 2.5 log10 CFU/g fewer organisms in spleen, and 1.8 to 2.5 log10 CFU/g fewer organisms in kidney, although differences were statistically significant only in spleen for daptomycin plus ceftriaxone and in kidney for daptomycin plus ertapenem. Drug exposures in rabbits were less than those achievable in humans, which may have limited the in vivo activity, particularly in vegetations. PMID:27090173

  14. Crystal-state structural analysis of two gamma-lactam-restricted analogs of Pro-Leu-Gly-NH2.

    PubMed

    Valle, G; Crisma, M; Toniolo, C; Yu, K L; Johnson, R L

    1989-03-01

    The crystal structures of two analogs of Pro-Leu-Gly-NH2 (1), containing a gamma-lactam conformational constraint in place of the -Leu-Gly- sequences, are described. The highly biologically active (S,R)-diastereomer 2a is semi-extended at the C-terminus, with the N-terminal Pro residue in the unusual "C5" conformation [psi 1 = -0.8(15) degrees] stabilized by a (peptide)N-H...N(amino) intramolecular H-bond [the N(3)...N(4) separation is 2.687(11)A]. Conversely, the N,N'-isopropylidene aminal trihydrate of the (S,S)-diastereomer 2b, compound 3, adopts a beta-bend conformation at the C-terminus, as already reported for 1. However, the backbone torsion angles [phi 2 = 57.4(4), psi 2 = -129.9(3) degrees; psi 3 = -92.3(4), phi 3 = 6.4(5) degrees] lie close to the values expected for the corner residues of an ideal type-II' beta-bend. A weak intramolecular 4----1 H-bond is seen between the Gly carboxyamide anti-NH and Pro C = O groups. In the newly formed 2,2,3,4-tetraalkyl-5-oxo-imidazolidin-1-yl moiety the psi 1 torsion angle is 12.9(4) degrees and the intramolecular N(3)...N(4) separation is 2.321(4)A. PMID:2565891

  15. Luteolin potentiates the effects of aminoglycoside and β-lactam antibiotics against methicillin-resistant Staphylococcus aureus in vitro

    PubMed Central

    JOUNG, DAE-KI; KANG, OK-HWA; SEO, YUN-SOO; ZHOU, TIAN; LEE, YOUNG-SEOB; HAN, SIN-HEE; MUN, SU-HYUN; KONG, RYONG; SONG, HO-JUN; SHIN, DONG-WON; KWON, DONG-YEUL

    2016-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) infection has become a serious clinical problem worldwide, and alternative natural or combination drug therapies are required for its treatment. The aim of the present study was to examined the antimicrobial activity of luteolin (LUT) against MRSA. Luteolin is a polyphenolic flavonoid compound with a wide spectrum of biological activities. The antimicrobial activities of LUT and the antibiotics ampicillin (AM), oxacillin (OX) and gentamicin (GT), used alone or in combination, were evaluated against five clinical MRSA isolates and two reference strains using a minimum inhibitory concentration (MIC) assay, MTT colorimetric assay, checkerboard dilution test and time-kill assay. The MIC of LUT against all strains was found to be 62.5 µg/ml. The combinations of LUT and antibiotics exhibited a synergistic effect against MRSA in the majority of cases, as determined by the checkerboard method. Time-kill curves revealed that a combination of LUT with AM, OX or GT significantly reduced bacterial counts, which dropped below the lowest detectable limit after 24 h. These results indicate that LUT potentiates the effects of β-lactam and aminoglycoside antibiotics against MRSA. PMID:27284353

  16. Isolation and characterization of a new strain of Achromobacter sp. with beta-lactam antibiotic acylase activity.

    PubMed

    Plhácková, K; Becka, S; Skrob, F; Kyslík, P

    2003-10-01

    A bacterial strain producing a beta-lactam antibiotic acylase, able to hydrolyze ampicillin to 6-aminopenicillanic acid more efficiently than penicillin G, was isolated from soil and characterized. The isolate was identified as Achromobacter sp. using the phenotypic characteristics, composition of cellular fatty acids and 16S rRNA gene sequence. The enzyme synthesis was fully induced by phenylacetic acid (PAA) at a concentration of 2 g l(-1). PAA at concentrations up to 12 g l(-1) had no negative effect on the specific activity of acylase and biomass production, but slowed down the specific growth rate. Benzoic or 4-hydroxyphenylacetic acids can also induce synthesis of the enzyme. The inducers were metabolized in all cases. Acylase activity in cell-free extracts was determined with various substrates; ampicillin, cephalexin and amoxicillin were hydrolyzed 1.5- and 2-times faster than penicillin G. A high stability of acylase activity was observed over a wide range of pH (5.0-8.5) and at temperatures above 55 degrees C. PMID:12827318

  17. The MazEF Toxin-Antitoxin System Alters the β-Lactam Susceptibility of Staphylococcus aureus.

    PubMed

    Schuster, Christopher F; Mechler, Lukas; Nolle, Nicoletta; Krismer, Bernhard; Zelder, Marc-Eric; Götz, Friedrich; Bertram, Ralph

    2015-01-01

    Toxin-antitoxin (TA) systems are genetic elements of prokaryotes which encode a stable toxin and an unstable antitoxin that can counteract toxicity. TA systems residing on plasmids are often involved in episomal maintenance whereas those on chromosomes can have multiple functions. The opportunistic pathogen Staphylococcus aureus possesses at least four different families of TA systems but their physiological roles are elusive. The chromosomal mazEF system encodes the RNase toxin MazF and the antitoxin MazE. In the light of ambiguity regarding the cleavage activity, we here verify that MazF specifically targets UACAU sequences in S. aureus in vivo. In a native strain background and under non-stress conditions, cleavage was observed in the absence or presence of mazE. Transcripts of spa (staphylococcal protein A) and rsbW (anti-σB factor) were cut, but translational reporter fusions indicated that protein levels of the encoded products were unaffected. Despite a comparable growth rate as the wild-type, an S. aureus mazEF deletion mutant was more susceptible to β-lactam antibiotics, which suggests that further genes, putatively involved in the antibiotic stress response or cell wall synthesis or turnover, are controlled by this TA system.

  18. Practical Management of Patients with a History of Immediate Hypersensitivity to Common non-Beta-Lactam Drugs.

    PubMed

    Macy, Eric

    2016-01-01

    Immediate hypersensitivity reactions to medications are among the most feared adverse drug reactions, because of their close association with anaphylaxis. This review discusses a practical management approach for patients with a history of an immediate hypersensitivity to a non-beta-lactam medication, where reexposure to the implicated, or similar, medication is clinically necessary. Mechanisms associated with severe immediate hypersensitivity reactions include IgE-mediated mast cell activation, complement-mediated mast cell activation, and direct mast cell activation. Immediate hypersensitivity reactions may also be mediated by vasodilators, other pharmacologic mechanisms, or be secondary to underlying patient-specific biochemical abnormalities such as endocrine tumors or chronic spontaneous urticaria. The key features in the reaction history and the biochemistry of the implicated medication are discussed. Most individuals with a history of immediate hypersensitivity to a medication, who require reuse of that drug, can be safely retreated with a therapeutic course of the implicated drug after a full-dose challenge, graded challenge, or desensitization, with or without premedication and/or any preliminary diagnostic testing, depending on the specific situation.

  19. The thioacetate-ω(γ-lactam carboxamide) HDAC inhibitor ST7612AA1 as HIV-1 latency reactivation agent.

    PubMed

    Badia, Roger; Grau, Judith; Riveira-Muñoz, Eva; Ballana, Ester; Giannini, Giuseppe; Esté, José A

    2015-11-01

    Antiretroviral therapy (ART) is unable to cure HIV infection. The ability of HIV to establish a subset of latent infected CD4(+) T cells, which remain undetectable to the immune system, becomes a major roadblock to achieve viral eradication. Histone deacetylase inhibitors (HDACi) have been shown to potently induce the reactivation of latent HIV. Here, we show that a new thiol-based HDACi, the thioacetate-ω(γ-lactam carboxamide) derivative ST7612AA1, is a potent inducer of HIV reactivation. We evaluated HIV reactivation activity of ST7612AA1 compared to panobinostat (PNB), romidepsin (RMD) and vorinostat (VOR) in cell culture models of HIV-1 latency, in latently infected primary CD4(+) T lymphocytes and in PBMCs from HIV(+) patients. ST7612AA1 potently induced HIV-1 reactivation at submicromolar concentrations with comparable potency to panobinostat or superior to vorinostat. The presence of known antiretrovirals did not affect ST7612AA1-induced reactivation and their activity was not affected by ST7612AA1. Cell proliferation and cell activation were not affected by ST7612AA1, or any other HDACi used. In conclusion, our results indicate that ST7612AA1 is a potent activator of latent HIV and that reactivation activity of ST7612AA1 is exerted without activation or proliferation of CD4(+) T cells. ST7612AA1 is a suitable candidate for further studies of HIV reactivation strategies and potential new therapies to eradicate the viral reservoirs. PMID:26348004

  20. Detection and reporting beta-lactam resistance phenotypes in Escherichia coli and Klebsiella pneumoniae: a multicenter proficiency study in Spain.

    PubMed

    Conejo, M Carmen; Mata, C; Navarro, F; Pascual, A

    2008-11-01

    The ability of 57 Spanish microbiology laboratories in detecting and reporting beta-lactam resistance phenotypes in Escherichia coli and Klebsiella pneumoniae was evaluated. Laboratories received 6 well-characterized isolates expressing the most widespread extended-spectrum beta-lactamases (ESBLs) in Spain (4 CTX-M type, 1 TEM type, and 1 SHV type), 3 isolates producing AmpC-type enzymes (2 plasmid mediated and 1 E. coli hyperproducing its chromosomal AmpC), and 3 quality control strains. Ninety-one percent of laboratories recognized all ESBL producers correctly, and therefore, low error rates were observed when testing cephalosporins and aztreonam. The highest error rates were observed with combinations of penicillin plus beta-lactamase inhibitor, although more than 60% of cases were due to the interpretation made by the microbiologists. Correct recognition of all AmpC beta-lactamase-producing strains occurred in only 47.4% of laboratories. These isolates were wrongly reported as ESBL producers and penicillinase hyperproducers in 7.6 % and 5.8% of cases, respectively. Detection of the AmpC-type phenotype by Spanish laboratories needs to be improved. PMID:18692340

  1. The MazEF Toxin-Antitoxin System Alters the β-Lactam Susceptibility of Staphylococcus aureus

    PubMed Central

    Schuster, Christopher F.; Mechler, Lukas; Nolle, Nicoletta; Krismer, Bernhard; Zelder, Marc-Eric; Götz, Friedrich; Bertram, Ralph

    2015-01-01

    Toxin-antitoxin (TA) systems are genetic elements of prokaryotes which encode a stable toxin and an unstable antitoxin that can counteract toxicity. TA systems residing on plasmids are often involved in episomal maintenance whereas those on chromosomes can have multiple functions. The opportunistic pathogen Staphylococcus aureus possesses at least four different families of TA systems but their physiological roles are elusive. The chromosomal mazEF system encodes the RNase toxin MazF and the antitoxin MazE. In the light of ambiguity regarding the cleavage activity, we here verify that MazF specifically targets UACAU sequences in S. aureus in vivo. In a native strain background and under non-stress conditions, cleavage was observed in the absence or presence of mazE. Transcripts of spa (staphylococcal protein A) and rsbW (anti-σB factor) were cut, but translational reporter fusions indicated that protein levels of the encoded products were unaffected. Despite a comparable growth rate as the wild-type, an S. aureus mazEF deletion mutant was more susceptible to β-lactam antibiotics, which suggests that further genes, putatively involved in the antibiotic stress response or cell wall synthesis or turnover, are controlled by this TA system. PMID:25965381

  2. Ceftriaxone, a beta-lactam antibiotic, attenuates relapse-like ethanol-drinking behavior in alcohol-preferring rats.

    PubMed

    Qrunfleh, Abeer M; Alazizi, Adnan; Sari, Youssef

    2013-06-01

    Relapse-like ethanol-drinking behavior depends on increased glutamate transmission in the mesocorticolimbic motive circuit. Extracellular glutamate is regulated by a number of glutamate transporters. Of these transporters, glutamate transporter 1 (GLT1) is responsible for the majority of extracellular glutamate uptake. We have recently reported that ceftriaxone (CEF) treatment (i.p.), a β-lactam antibiotic known to elevate GTL1 expression, reduced ethanol intake in male alcohol-preferring (P) rats. We investigated here whether CEF treatment attenuates relapse-like ethanol-drinking behavior. P rats were exposed to free choice of 15% and 30% ethanol for 5 weeks and treated with CEF (50 and 100 mg/kg, i.p.) during the last 5 days of the 2-week deprivation period. Rats treated with CEF during the deprivation period showed a reduction in ethanol intake compared with saline-treated rats upon re-exposure to ethanol; this effect persisted for 9 days. Moreover, CEF-mediated attenuation in relapse to ethanol-drinking behavior was associated with upregulation of GLT1 level in prefrontal cortex and nucleus accumbens core. GLT1 upregulation was revealed only at the higher dose of CEF. In addition, CEF has no effect on relapse-like sucrose-drinking behavior. These findings suggest that ceftriaxone might be used as a potential therapeutic treatment for the attenuation of relapse-like ethanol-drinking behavior.

  3. Ceftazidime/Avibactam and Ceftolozane/Tazobactam: Second-generation β-Lactam/β-Lactamase Inhibitor Combinations.

    PubMed

    van Duin, David; Bonomo, Robert A

    2016-07-15

    Ceftolozane/tazobactam and ceftazidime/avibactam are 2 novel β-lactam/β-lactamase combination antibiotics. The antimicrobial spectrum of activity of these antibiotics includes multidrug-resistant (MDR) gram-negative bacteria (GNB), including Pseudomonas aeruginosa. Ceftazidime/avibactam is also active against carbapenem-resistant Enterobacteriaceae that produce Klebsiella pneumoniae carbapenemases. However, avibactam does not inactivate metallo-β-lactamases such as New Delhi metallo-β-lactamases. Both ceftolozane/tazobactam and ceftazidime/avibactam are only available as intravenous formulations and are dosed 3 times daily in patients with normal renal function. Clinical trials showed noninferiority to comparators of both agents when used in the treatment of complicated urinary tract infections and complicated intra-abdominal infections (when used with metronidazole). Results from pneumonia studies have not yet been reported. In summary, ceftolozane/tazobactam and ceftazidime/avibactam are 2 new second-generation cephalosporin/β-lactamase inhibitor combinations. After appropriate trials are conducted, they may prove useful in the treatment of MDR GNB infections. Antimicrobial stewardship will be essential to preserve the activity of these agents.

  4. Electric stimulations mediated beta lactam resistance reversal and correlation with growth dynamics of community acquired methicillin resistant Staphylococcus aureus.

    PubMed

    Kainthola, Anup; Uniyal, Akshat; Srivastava, Nidhi; Bhatt, Ajay B

    2015-08-01

    The community associated methicillin resistant Staphylococcus aureus (CA-MRSA) is a serious issue of public health. Here, we conducted an experimental approach to determine: (i) the optimal significant stimulation range of electrical current for effective checking of CA-MRSA growth; (ii) the effect of electrical stimulations on methicillin susceptibility and possible beta lactam resistance reversal; and (iii) the variation in the level of ATP as function of exposure to electric current. An 8 chambered electrical system was developed for DC flow in control and test sets, with and without drug (oxacillin 4 mg/ml). Measurement of growth by CFU/ml and spectrometry, susceptibility and ATP levels were calculated and interpreted. Linear pattern in reduction of ATP was observed with respect to the intensity of electric current (EC) and an enhanced inhibitory effect was explicit with 1000 microampere (μA) with 30 min exposure. At 4000 μA exposure to DC at 180 min and in combination of drug (μA+D), the growth of CA-MRSA was substantially checked to 0.23 absorbance in comparison to current without drug and the effect of DC electrical current to the culture showed that 10 μA, 100 μA and 4000 μA current exposure in combination of oxacillin (μA+D), markedly reduced the CFU to an average of 256.4. ATP level was linearly reduced with exposure to EC.

  5. Resistance to ceftriaxone and other beta-lactams in bacteria isolated in the community. The Vigil'Roc Study Group.

    PubMed Central

    Goldstein, F W; Péan, Y; Gertner, J

    1995-01-01

    The incidence of bacterial species and their susceptibilities to ceftriaxone and other beta-lactams from patients with community-acquired infections were evaluated in a multicenter study over a 4-month period. A total of 5,768 bacterial isolates were classified according to whether the patient had been previously hospitalized or had received antibiotic treatment. The most relevant findings were the presence of 33.8% penicillin-resistant Streptococcus pneumoniae isolates, 25% beta-lactamase-producing Haemophilus influenzae isolates, and 36.4% amoxicillin-resistant Escherichia coli isolates. All of these bacteria were fully susceptible to ceftriaxone. Nosocomial multiply-resistant bacteria, and particularly methicillin-resistant S. aureus, were found, as expected, at a higher frequency among previously hospitalized patients. However, such bacteria may be present in the community; their incidence is high in particular clinical settings, and such bacteria should be considered when one is choosing a first-line therapy for the treatment of severe infections. PMID:8585736

  6. Immobilization on macroporous resin makes E. coli RutB a robust catalyst for production of (-) Vince lactam.

    PubMed

    Wang, Jianjun; Zhu, Junge; Wu, Sheng

    2015-06-01

    A novel (+) γ-lactamase gene (rutB) was cloned from Escherichia coli JM109 and expressed in E. coli BL21 (DE3), and the recombinant protein was characterized. The optimal conditions for the enzyme were pH 7.0 and temperature 30 °C, which indicated that it was a mesophilic protein. The free purified enzyme was deactivated when incubated at 50 °C for 30 min. However, the k cat value of RutB at its optimal temperature was about 2.5 times that of the archaeal enzyme from Sulfolobus sofataricus at its optimal temperature (85 °C). After immobilization on macroporous resin using glutaraldehyde cross-linkage, the thermostability of the crude enzyme was greatly enhanced and the deactivating temperature was raised to 70 °C. After immobilization, the minimal substrate inhibition concentration for RutB also improved from 0.75 to 1.5 M. The optimal concentrations of immobilized enzyme and substrate were determined to be 250 mg/ml and 1.5 M, when the initial reaction velocity was the response variable in batch transformations. This immobilization of RutB on macroporous resins provides another feasible approach for the preparation of optically active Vince lactam. As a member of the isochorismatase superfamily, RutB was demonstrated to be another typical γ-lactamase that showed catalytic promiscuity.

  7. Predictability of Phenotype in Relation to Common β-Lactam Resistance Mechanisms in Escherichia coli and Klebsiella pneumoniae.

    PubMed

    Agyekum, Alex; Fajardo-Lubián, Alicia; Ai, Xiaoman; Ginn, Andrew N; Zong, Zhiyong; Guo, Xuejun; Turnidge, John; Partridge, Sally R; Iredell, Jonathan R

    2016-05-01

    The minimal concentration of antibiotic required to inhibit the growth of different isolates of a given species with no acquired resistance mechanisms has a normal distribution. We have previously shown that the presence or absence of transmissible antibiotic resistance genes has excellent predictive power for phenotype. In this study, we analyzed the distribution of six β-lactam antibiotic susceptibility phenotypes associated with commonly acquired resistance genes in Enterobacteriaceae in Sydney, Australia. Escherichia coli (n = 200) and Klebsiella pneumoniae (n = 178) clinical isolates, with relevant transmissible resistance genes (blaTEM, n = 33; plasmid AmpC, n = 69; extended-spectrum β-lactamase [ESBL], n = 116; and carbapenemase, n = 100), were characterized. A group of 60 isolates with no phenotypic resistance to any antibiotics tested and carrying none of the important β-lactamase genes served as comparators. The MICs for all drug-bacterium combinations had a normal distribution, varying only in the presence of additional genes relevant to the phenotype or, for ertapenem resistance in K. pneumoniae, with a loss or change in the outer membrane porin protein OmpK36. We demonstrated mutations in ompK36 or absence of OmpK36 in all isolates in which reduced susceptibility to ertapenem (MIC, >1 mg/liter) was evident. Ertapenem nonsusceptibility in K. pneumoniae was most common in the context of an OmpK36 variant with an ESBL or AmpC gene. Surveillance strategies to define appropriate antimicrobial therapies should include genotype-phenotype relationships for all major transmissible resistance genes and the characterization of mutations in relevant porins in organisms, like K. pneumoniae. PMID:26912748

  8. Beta Lactams Antibiotic Ceftriaxone Modulates Seizures, Oxidative Stress and Connexin 43 Expression in Hippocampus of Pentylenetetrazole Kindled Rats

    PubMed Central

    Hussein, Abdelaziz M.; Ghalwash, Mohammed; Magdy, Khaled; Abulseoud, Osama A.

    2016-01-01

    Background and Purpose: This study aimed to investigate the effect of ceftriaxone on oxidative stress and gap junction protein (connexin 43, Cx-43) expression in pentylenetetrazole (PTZ) induced kindling model. Methods: Twenty four Sprague dawely rats were divided into 3 equal groups (a) normal group: normal rats. (b) PTZ kindled group: received PTZ at the dose of 50 mg/kg via intraperitoneal injection (i.p.) every other day for 2 weeks (c) ceftriaxone treated group: received ceftriaxone at the dose 200 mg\\kg/12 hrs via i.p. injection daily from the 6th dose of PTZ for 3 days. Racine score, latency before beginning the first myoclonic jerk and duration of the jerks used as parameters of behavioral assessment. Immunohistopathological study for Cx-43 expression in hippocampus and measurement of markers of oxidative stress (malondialdehyde [MDA], low reduced glutathione [GSH] and catalase [CAT]) in hippocampal neurons were done. Results: PTZ kindling was associated with behavioral changes (in the form high stage of Racine score, long seizure duration and short latency for the first jerk), enhanced oxidative stress state (as demonstrated by high MDA, low GSH and CAT) and up regulation of Cx43 in hippocampal regions. While, ceftriaxone treatment ameliorated, significantly, PTZ-induced convulsions and caused significant improvement in oxidative stress markers and Cx-43 expression in hippocamal regions (p < 0.05). Conclusions: These findings support the anticonvulsive effects of some beta-lactams antibiotics which could offer a possible contributor in the basic treatment of temporal lobe epilepsy. This effect might be due to reduction of oxidative stress and Cx43 expression. PMID:27390674

  9. Haemophilus influenzae with Non-Beta-Lactamase-Mediated Beta-Lactam Resistance: Easy To Find but Hard To Categorize

    PubMed Central

    Lia, Astrid; Hannisdal, Anja; Tveten, Yngvar; Matuschek, Erika; Kahlmeter, Gunnar; Kristiansen, Bjørn-Erik

    2015-01-01

    Haemophilus influenzae is a major pathogen, and beta-lactams are first-line drugs. Resistance due to altered penicillin-binding protein 3 (rPBP3) is frequent, and susceptibility testing of such strains is challenging. A collection of 154 beta-lactamase-negative isolates with a large proportion of rPBP3 (67.5%) was used to evaluate and compare Etest (Haemophilus test medium [HTM]) and disk diffusion (EUCAST method) for categorization of susceptibility to aminopenicillins and cefuroxime, using MICs generated with broth (HTM) microdilution and clinical breakpoints from CLSI and EUCAST as the gold standards. In addition, the proficiency of nine disks in screening for the rPBP3 genotype (N526K positive) was evaluated. By Etest, both essential and categorical agreement were generally poor (<70%), with high very major errors (VME) (CLSI, 13.0%; EUCAST, 34.3%) and falsely susceptible rates (FSR) (CLSI, 87.0%; EUCAST, 88.3%) for ampicillin. Ampicillin (2 μg) with adjusted (+2 mm) zone breakpoints was superior to Etest for categorization of susceptibility to ampicillin (agreement, 74.0%; VME, 11.0%; FSR, 28.3%). Conversely, Etest was superior to 30 μg cefuroxime for categorization of susceptibility to cefuroxime (agreement, 57.1% versus 60.4%; VME, 2.6% versus 9.7%; FSR, 7.1% versus 26.8%). Benzylpenicillin (1 unit) (EUCAST screening disk) and cefuroxime (5 μg) identified rPBP3 isolates with highest accuracies (95.5% and 92.2%, respectively). In conclusion, disk screening reliably detects rPBP3 H. influenzae, but false ampicillin susceptibility is frequent with routine methods. We suggest adding a comment recommending high-dose aminopenicillin therapy or the use of other agents for severe infections with screening-positive isolates that are susceptible to aminopenicillins by gradient or disk diffusion. PMID:26354813

  10. Ampicillin-Sulbactam and Amoxicillin-Clavulanate Susceptibility Testing of Escherichia coli Isolates with Different β-Lactam Resistance Phenotypes

    PubMed Central

    Oliver, Antonio; Pérez-Vázquez, María; Martínez-Ferrer, Manuel; Baquero, Fernando; de Rafael, Luis; Cantón, Rafael

    1999-01-01

    The activities of ampicillin-sulbactam and amoxicillin-clavulanate were studied with 100 selected clinical Escherichia coli isolates with different β-lactam susceptibility phenotypes by standard agar dilution and disk diffusion techniques and with a commercial microdilution system (PASCO). A fixed ratio (2:1) and a fixed concentration (clavulanate, 2 and 4 μg/ml; sulbactam, 8 μg/ml) were used in the agar dilution technique. The resistance frequencies for amoxicillin-clavulanate with different techniques were as follows: fixed ratio agar dilution, 12%; fixed concentration 4-μg/ml agar dilution, 17%; fixed ratio microdilution, 9%; and disk diffusion, 9%. Marked discrepancies were found when these results were compared with those obtained with ampicillin-sulbactam (26 to 52% resistance), showing that susceptibility to amoxicillin-clavulanic acid cannot be predicted by testing the isolate against ampicillin-sulbactam. Interestingly, the discrimination between susceptible and intermediate isolates was better achieved with 4 μg of clavulanate per ml than with the fixed ratio. In contrast, amoxicillin susceptibility was not sufficiently restored when 2 μg of clavulanate per ml was used, particularly in moderate (mean β-lactamase activity, 50.8 mU/mg of protein) and high-level (215 mU/mg) TEM-1 β-lactamase producer isolates. Four micrograms of clavulanate per milliliter could be a reasonable alternative to the 2:1 fixed ratio, because most high-level β-lactamase-hyperproducing isolates would be categorized as nonsusceptible, and low- and moderate-level β-lactamase-producing isolates would be categorized as nonresistant. This approach cannot be applied to sulbactam, either with the fixed 2:1 ratio or with the 8-μg/ml fixed concentration, because many low-level β-lactamase-producing isolates would be classified in the resistant category. These findings call for a review of breakpoints for β-lactam–β-lactamase inhibitor combinations. PMID:10103192

  11. Haemophilus influenzae with Non-Beta-Lactamase-Mediated Beta-Lactam Resistance: Easy To Find but Hard To Categorize.

    PubMed

    Skaare, Dagfinn; Lia, Astrid; Hannisdal, Anja; Tveten, Yngvar; Matuschek, Erika; Kahlmeter, Gunnar; Kristiansen, Bjørn-Erik

    2015-11-01

    Haemophilus influenzae is a major pathogen, and beta-lactams are first-line drugs. Resistance due to altered penicillin-binding protein 3 (rPBP3) is frequent, and susceptibility testing of such strains is challenging. A collection of 154 beta-lactamase-negative isolates with a large proportion of rPBP3 (67.5%) was used to evaluate and compare Etest (Haemophilus test medium [HTM]) and disk diffusion (EUCAST method) for categorization of susceptibility to aminopenicillins and cefuroxime, using MICs generated with broth (HTM) microdilution and clinical breakpoints from CLSI and EUCAST as the gold standards. In addition, the proficiency of nine disks in screening for the rPBP3 genotype (N526K positive) was evaluated. By Etest, both essential and categorical agreement were generally poor (<70%), with high very major errors (VME) (CLSI, 13.0%; EUCAST, 34.3%) and falsely susceptible rates (FSR) (CLSI, 87.0%; EUCAST, 88.3%) for ampicillin. Ampicillin (2 μg) with adjusted (+2 mm) zone breakpoints was superior to Etest for categorization of susceptibility to ampicillin (agreement, 74.0%; VME, 11.0%; FSR, 28.3%). Conversely, Etest was superior to 30 μg cefuroxime for categorization of susceptibility to cefuroxime (agreement, 57.1% versus 60.4%; VME, 2.6% versus 9.7%; FSR, 7.1% versus 26.8%). Benzylpenicillin (1 unit) (EUCAST screening disk) and cefuroxime (5 μg) identified rPBP3 isolates with highest accuracies (95.5% and 92.2%, respectively). In conclusion, disk screening reliably detects rPBP3 H. influenzae, but false ampicillin susceptibility is frequent with routine methods. We suggest adding a comment recommending high-dose aminopenicillin therapy or the use of other agents for severe infections with screening-positive isolates that are susceptible to aminopenicillins by gradient or disk diffusion. PMID:26354813

  12. Design and synthesis of pyrrolidine-5,5-trans-lactams (5-oxohexahydropyrrolo[3,2-b]pyrroles) as novel mechanism-based inhibitors of human cytomegalovirus protease. 2. Potency and chirality.

    PubMed

    Borthwick, Alan D; Crame, Andrew J; Ertl, Peter F; Exall, Anne M; Haley, Terry M; Hart, Graham J; Mason, Andrew M; Pennell, Andrew M K; Singh, Onkar M P; Weingarten, Gordon G; Woolven, James M

    2002-01-01

    The stereospecific synthesis of a series of alpha-methylpyrrolidine-5,5-trans-lactam inhibitors of human cytomegalovirus (HCMV) protease is described. Examination of the SAR in this series has defined the size and chirality of the alpha-substituent, optimized the acyl substituent on the lactam nitrogen, and defined the steric constraint of this functionality. The SAR of the functionality on the pyrrolidine nitrogen of the trans-lactam has been investigated, and this has led to the discovery of potent serine protease inhibitors that are highly selective for the viral enzyme over the mammalian enzymes elastase, thrombin, and acetylcholine esterase. The mechanism of action of our lead compounds has been established by mass spectrometry, and enzymatic degradation of HCMV deltaAla protease acylated with these inhibitors showed that Ser 132 is the active site nucleophile. The crystal structure of HCMV protease was obtained and used to model the conformationally restricted, chiral (S)-proline-alpha-methyl-5,5-trans-lactams into the active site groove of the enzyme, enabling us to direct and rationalize the SAR in this series. The activity against HCMV deltaAla protease is the greatest with inhibitors based on the dansyl-(S)-proline alpha-methyl-5,5-trans-lactam template, which have low nanomolar activity against the viral enzyme.

  13. Evaluation of the Charm maximum residue limit β-lactam and tetracycline test for the detection of antibiotics in ewe and goat milk.

    PubMed

    Beltrán, M C; Romero, T; Althaus, R L; Molina, M P

    2013-05-01

    The Charm maximum residue limit β-lactam and tetracycline test (Charm MRL BLTET; Charm Sciences Inc., Lawrence, MA) is an immunoreceptor assay utilizing Rapid One-Step Assay lateral flow technology that detects β-lactam or tetracycline drugs in raw commingled cow milk at or below European Union maximum residue levels (EU-MRL). The Charm MRL BLTET test procedure was recently modified (dilution in buffer and longer incubation) by the manufacturers to be used with raw ewe and goat milk. To assess the Charm MRL BLTET test for the detection of β-lactams and tetracyclines in milk of small ruminants, an evaluation study was performed at Instituto de Ciencia y Tecnologia Animal of Universitat Politècnica de València (Spain). The test specificity and detection capability (CCβ) were studied following Commission Decision 2002/657/EC. Specificity results obtained in this study were optimal for individual milk free of antimicrobials from ewes (99.2% for β-lactams and 100% for tetracyclines) and goats (97.9% for β-lactams and 100% for tetracyclines) along the entire lactation period regardless of whether the results were visually or instrumentally interpreted. Moreover, no positive results were obtained when a relatively high concentration of different substances belonging to antimicrobial families other than β-lactams and tetracyclines were present in ewe and goat milk. For both types of milk, the CCβ calculated was lower or equal to EU-MRL for amoxicillin (4 µg/kg), ampicillin (4 µg/kg), benzylpenicillin (≤ 2 µg/kg), dicloxacillin (30 µg/kg), oxacillin (30 µg/kg), cefacetrile (≤ 63 µg/kg), cefalonium (≤ 10 µg/kg), cefapirin (≤ 30 µg/kg), desacetylcefapirin (≤ 30 µg/kg), cefazolin (≤ 25 µg/kg), cefoperazone (≤ 25 µg/kg), cefquinome (20 µg/kg), ceftiofur (≤ 50 µg/kg), desfuroylceftiofur (≤ 50µg/kg), and cephalexin (≤ 50 µg/kg). However, this test could neither detect cloxacillin nor nafcillin at or below EU-MRL (CCβ >30 µg/kg). The

  14. Combinations of β-lactam or aminoglycoside antibiotics with plectasin are synergistic against methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

    PubMed

    Hu, Yanmin; Liu, Alexander; Vaudrey, James; Vaiciunaite, Brigita; Moigboi, Christiana; McTavish, Sharla M; Kearns, Angela; Coates, Anthony

    2015-01-01

    Bacterial infections remain the leading killer worldwide which is worsened by the continuous emergence of antibiotic resistance. In particular, methicillin-sensitive (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) are prevalent and the latter can be difficult to treat. The traditional strategy of novel therapeutic drug development inevitably leads to emergence of resistant strains, rendering the new drugs ineffective. Therefore, rejuvenating the therapeutic potentials of existing antibiotics offers an attractive novel strategy. Plectasin, a defensin antimicrobial peptide, potentiates the activities of other antibiotics such as β-lactams, aminoglycosides and glycopeptides against MSSA and MRSA. We performed in vitro and in vivo investigations to test against genetically diverse clinical isolates of MSSA (n = 101) and MRSA (n = 115). Minimum inhibitory concentrations (MIC) were determined by the broth microdilution method. The effects of combining plectasin with β-lactams, aminoglycosides and glycopeptides were examined using the chequerboard method and time kill curves. A murine neutropenic thigh model and a murine peritoneal infection model were used to test the effect of combination in vivo. Determined by factional inhibitory concentration index (FICI), plectasin in combination with aminoglycosides (gentamicin, neomycin or amikacin) displayed synergistic effects in 76-78% of MSSA and MRSA. A similar synergistic response was observed when plectasin was combined with β-lactams (penicillin, amoxicillin or flucloxacillin) in 87-89% of MSSA and MRSA. Interestingly, no such interaction was observed when plectasin was paired with vancomycin. Time kill analysis also demonstrated significant synergistic activities when plectasin was combined with amoxicillin, gentamicin or neomycin. In the murine models, plectasin at doses as low as 8 mg/kg augmented the activities of amoxicillin and gentamicin in successful treatment of MSSA and MRSA infections. We

  15. Combinations of β-Lactam or Aminoglycoside Antibiotics with Plectasin Are Synergistic against Methicillin-Sensitive and Methicillin-Resistant Staphylococcus aureus

    PubMed Central

    Hu, Yanmin; Liu, Alexander; Vaudrey, James; Vaiciunaite, Brigita; Moigboi, Christiana; McTavish, Sharla M.; Kearns, Angela; Coates, Anthony

    2015-01-01

    Bacterial infections remain the leading killer worldwide which is worsened by the continuous emergence of antibiotic resistance. In particular, methicillin-sensitive (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) are prevalent and the latter can be difficult to treat. The traditional strategy of novel therapeutic drug development inevitably leads to emergence of resistant strains, rendering the new drugs ineffective. Therefore, rejuvenating the therapeutic potentials of existing antibiotics offers an attractive novel strategy. Plectasin, a defensin antimicrobial peptide, potentiates the activities of other antibiotics such as β-lactams, aminoglycosides and glycopeptides against MSSA and MRSA. We performed in vitro and in vivo investigations to test against genetically diverse clinical isolates of MSSA (n = 101) and MRSA (n = 115). Minimum inhibitory concentrations (MIC) were determined by the broth microdilution method. The effects of combining plectasin with β-lactams, aminoglycosides and glycopeptides were examined using the chequerboard method and time kill curves. A murine neutropenic thigh model and a murine peritoneal infection model were used to test the effect of combination in vivo. Determined by factional inhibitory concentration index (FICI), plectasin in combination with aminoglycosides (gentamicin, neomycin or amikacin) displayed synergistic effects in 76-78% of MSSA and MRSA. A similar synergistic response was observed when plectasin was combined with β-lactams (penicillin, amoxicillin or flucloxacillin) in 87–89% of MSSA and MRSA. Interestingly, no such interaction was observed when plectasin was paired with vancomycin. Time kill analysis also demonstrated significant synergistic activities when plectasin was combined with amoxicillin, gentamicin or neomycin. In the murine models, plectasin at doses as low as 8 mg/kg augmented the activities of amoxicillin and gentamicin in successful treatment of MSSA and MRSA infections. We

  16. Beta-lactam antibiotic-induced platelet dysfunction: Evidence for irreversible inhibition of platelet activation in vitro and in vivo after prolonged exposure to penicillin

    SciTech Connect

    Burroughs, S.F.; Johnson, G.J. )

    1990-04-01

    beta-Lactam antibiotics cause platelet dysfunction with bleeding complications. Previous in vitro studies documented reversible inhibition of agonist-receptor interaction. This mechanism is inadequate to explain the effect of beta-lactam antibiotics in vivo. Platelet function does not return to normal immediately after drug treatment, implying irreversible inhibition of platelet function. We report here evidence of irreversible platelet functional and biochemical abnormalities after in vitro and in vivo exposure to beta-lactam antibiotics. Irreversible binding of (14C)-penicillin (Pen) occurred in vitro. After 24 hours' in vitro incubation with 10 to 20 mmol/L Pen, or ex vivo after antibiotic treatment, irreversible functional impairment occurred; but no irreversible inhibition of alpha 2 adrenergic receptors, measured with (3H)-yohimbine, or high-affinity thromboxane A2/prostaglandin H2 (TXA2/PGH2) receptors, measured with agonist (3H)-U46619 and antagonist (3H)-SQ29548, occurred. However, low-affinity platelet TXA2/PGH2 receptors were decreased 40% after Pen exposure in vitro or in vivo, indicating irreversible membrane alteration. Two postreceptor biochemical events were irreversibly inhibited in platelets incubated with Pen for 24 hours in vitro or ex vivo after antibiotic treatment. Thromboxane synthesis was inhibited 28.3% to 81.7%. Agonist-induced rises in cytosolic calcium ((Ca2+)i) were inhibited 40.1% to 67.5% in vitro and 26.6% to 52.2% ex vivo. Therefore, Pen binds to platelets after prolonged exposure, resulting in irreversible dysfunction attributable to inhibition of TXA2 synthesis and impairment of the rise in (Ca2+)i. The loss of low-affinity TXA2/PGH2 receptors suggests that the primary site of action of these drugs is on the platelet membrane.

  17. Hospital outbreak of Klebsiella pneumoniae resistant to broad-spectrum cephalosporins and beta-lactam-beta-lactamase inhibitor combinations by hyperproduction of SHV-5 beta-lactamase.

    PubMed Central

    French, G L; Shannon, K P; Simmons, N

    1996-01-01

    An aminoglycoside- and ceftazidime-resistant strain of Klebsiella pneumoniae K2 producing the extended-spectrum beta-lactamase SHV-5 infected or colonized 14 pediatric patients at Guy's Hospital. The patients were mostly neonates recovering from cardiac surgery for congenital defects. The organism was also isolated from a nurse and from the father of one of the children. Four patients had septicemia, and two septicemic neonates with postoperative renal failure died. Aminoglycoside and cephalosporin resistance transferred to Escherichia coli in vitro on a 160-kb plasmid, and a similar resistant E. coli strain was isolated from the stools of one of the affected children. The epidemic organism colonized the bowel and skin and was probably transmitted via staff hands. Five wards were involved because of extensive patient movements. The outbreak was controlled by patient isolation and attention to handwashing. All of the isolates of the outbreak strain were identical by phage typing, ribotyping, plasmid profiling, and biochemical and serological testing, but they varied in their production of SHV-5. Some isolates produced normal amounts of SHV-5 and were susceptible to beta-lactam-beta-lactamase inhibitor combinations. Others, including the single isolate of multiresistant E. coli, produced up to five times as much enzyme as "normal" isolates. This hyperproduction resulted in increased resistance to several penicillins and cephalosporins and to the beta-lactam-beta-lactamase inhibitor combinations amoxicillin-clavulanic acid, ampicillin-sulbactam, piperacillin-tazobactam, and ceftazidime-clavulanic acid. The hyperproduction of SHV-5 by K. pneumoniae and E. coli seen in this outbreak suggests that beta-lactam-beta-lactamase inhibitor combinations may be unreliable for the treatment of organisms producing extended-spectrum beta-lactamases. PMID:8789016

  18. Plasmid-mediated AmpC-type beta-lactamase isolated from Klebsiella pneumoniae confers resistance to broad-spectrum beta-lactams, including moxalactam.

    PubMed Central

    Horii, T; Arakawa, Y; Ohta, M; Ichiyama, S; Wacharotayankun, R; Kato, N

    1993-01-01

    Klebsiella pneumoniae NU2936 was isolated from a patient and was found to produce a plasmid-encoded beta-lactamase (MOX-1) which conferred resistance to broad spectrum beta-lactams, including moxalactam, flomoxef, ceftizoxime, cefotaxime, and ceftazidime. Resistance could be transferred from K. pneumoniae NU2936 to Escherichia coli CSH2 by conjugation with a transfer frequency of 5 x 10(-7). The structural gene of MOX-1 (blaMOX-1) was cloned and expressed in E. coli HB101. The MIC of moxalactam for E. coli HB101 producing MOX-1 was > 512 micrograms/ml. The apparent molecular mass and pI of this enzyme were calculated to be 38 kDa and 8.9, respectively. Hg2+ and Cu2+ failed to block enzyme activity, and the presence of EDTA in the reaction buffer did not reduce the enzyme activity. However, clavulanate and cloxacillin, serine beta-lactamase inhibitors, inhibited the enzyme activity competitively (Kis = 5.60 and 0.35 microM, respectively). The kinetic study of MOX-1 suggested that it effectively hydrolyzed broad-spectrum beta-lactams. A hybridization study confirmed that blaMOX-1 is encoded on a large resident plasmid (pRMOX1; 180 kb) of strain NU2936. By deletion analysis, the functional region was localized within a 1.2-kb region of the plasmid. By amino acid sequencing, 18 of 33 amino acid residues at the N terminus of MOX-1 were found to be identical to those of Pseudomonas aeruginosa AmpC. These findings suggest that MOX-1 is a plasmid-mediated AmpC-type beta-lactamase that provides enteric bacteria resistance to broad-spectrum beta-lactams, including moxalactam. Images PMID:8517725

  19. A New Metabolite with a Unique 4-Pyranone-γ-Lactam-1,4-Thiazine Moiety from a Hawaiian-Plant Associated Fungus.

    PubMed

    Li, Chun-Shun; Ding, Yuanqing; Yang, Bao-Jun; Miklossy, Gabriella; Yin, Hong-Quan; Walker, Larry A; Turkson, James; Cao, Shugeng

    2015-07-17

    An endophytic fungus Paraphaeosphaeria neglecta FT462 isolated from the Hawaiian-plant Lycopodiella cernua (L.) Pic. Serm produced one unusual compound (1, paraphaeosphaeride A) with the 4-pyranone-γ-lactam-1,4-thiazine moiety, along with two new compounds (2 and 3, paraphaeosphaerides B and C, respectively) and the known compound (4). Compounds 1-3 were characterized by NMR and MS spectroscopic analysis. The absolute configuration of the 3-position of compound 1 was determined as S by electronic circular dichroism (ECD) calculations. Compound 3 also showed STAT3 inhibition at 10 μM. PMID:26107089

  20. Synthesis, structure, optical properties, antifungal and antibacterial activities of 2-(1-oxo-1H-2,3-dihydroisoindol-2-yl)-3-imidazolyl-L-lactamic acid

    NASA Astrophysics Data System (ADS)

    Jia, Ting; Zhang, Wei-Long; Chen, Yun; Cai, Shuang-Lian; Yi, Hai-Bo

    2013-10-01

    2-(1-oxo-1H-2,3-dihydroisoindol-2-yl)-3-imidazolyl-L-lactamic acid has been prepared conveniently by the condensation reaction of o-phthalaldehyde (OPA) with L-Histidine, and its single crystal structure has been characterized by X-ray crystallography method. The in vitro antifungal and antibacterial activities of the compound were investigated with the representative strains of Candida albicans, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. Its luminescent and nonlinear optical properties have also been investigated. Second-harmonic-generation (SHG) measurements indicate that compound 1 displays a weak SHG response of about 0.75 times that of KH2PO4.

  1. Preincubation of pneumococci with beta-lactams alone or combined with levofloxacin prevents quinolone-induced resistance without increasing intracellular levels of levofloxacin.

    PubMed

    Cottagnoud, Philippe; Johnson, Maggie; Cottagnoud, Marianne; Piddock, Laura

    2005-08-01

    Preincubation of pneumococci with sub-MIC concentrations of ceftriaxone (1/16x MIC), cefotaxime (1/8x MIC), and meropenem (1/4x MIC) alone or combined with levofloxacin (1/8x MIC) over 6 h prevents the emergence of levofloxacin-resistant mutants after 96 h of incubation but does not affect the intracellular accumulation of levofloxacin in two penicillin-resistant pneumococcal strains, suggesting a link between the mechanism of action of beta-lactams and the emergence of quinolone-induced resistance in pneumococci.

  2. Tetrahydro-β-carboline-based spirocyclic lactam as type II' β-turn: application to the synthesis and biological evaluation of somatostatine mimetics.

    PubMed

    Lesma, Giordano; Cecchi, Roberto; Cagnotto, Alfredo; Gobbi, Marco; Meneghetti, Fiorella; Musolino, Manuele; Sacchetti, Alessandro; Silvani, Alessandra

    2013-03-15

    The synthesis of novel spirocyclic lactams, embodying D-tryptophan (Trp) amino acid as the central core and acting as peptidomimetics, is presented. It relies on the strategic combination of Seebach's self-reproduction of chirality chemistry and Pictet-Spengler condensation as key steps. Investigation of the conformational behavior by molecular modeling, X-ray crystallography, and NMR and IR spectroscopies suggests very stable and highly predictable type II' β-turn conformations for all compounds. Relying on this feature, we also pursued their application to two potential mimetics of the hormone somatostatin, a pharmaceutically relevant natural peptide, which contains a Trp-based type II' β-turn pharmacophore.

  3. A novel β-lactam derivative, albactam from the flowers of Albizia lebbeck with platelets anti-aggregatory activity in vitro.

    PubMed

    El-Gamal, Ali Ali; Abd-El-Halim, Mohamed Farag; Kalil, Ashraf Taha; Basudan, Omer Ahmed; Al-Rehaily, Adnan Jathlan; Ahmad, Mohamed Shamim; El-Tahir, Kamal Hussin; Al-Massarani, Shaza Mohamed; Abdel-Mageed, Wael Moustafa

    2015-03-01

    A novel β-lactam derivative, albactam, was isolated from the alcoholic extract of the flowers of Albizia lebbeck. It showed a significant anti-aggregatory activity against adenosine diphosphate and arachidonic acid induced guinea-pigs' platelets aggregation in vitro. Six more known compounds were also isolated and fully characterized by measuring 1D and 2D NMR, two of them are the triterpenes β-amyrin and 11α, 12α-oxidotaraxerol, two ceramide derivatives and two flavonoids, kampferol 3-O-rutinoside and rutin. PMID:25796149

  4. Oritavancin Combinations with β-Lactams against Multidrug-Resistant Staphylococcus aureus and Vancomycin-Resistant Enterococci.

    PubMed

    Smith, Jordan R; Yim, Juwon; Raut, Animesh; Rybak, Michael J

    2016-04-01

    Oritavancin possesses activity against vancomycin-resistant enterococci (VRE) and methicillin-resistantStaphylococcus aureus(MRSA).In vitrodata suggest synergy between beta-lactams (BLs) and vancomycin or daptomycin, agents similar to oritavancin. We evaluated the activities of BLs combined with oritavancin against MRSA and VRE. Oritavancin MICs were determined for 30 strains, 5 each of MRSA, daptomycin-nonsusceptible (DNS) MRSA, vancomycin-intermediate MRSA (VISA), heteroresistant VISA (hVISA), vancomycin-resistantEnterococcus faecalis, and vancomycin-resistantEnterococcus faecium Oritavancin MICs were determined in the presence of subinhibitory concentrations of BLs. Oritavancin combined with ceftaroline, cefazolin, or nafcillin was evaluated for lethal synergy against MRSA, and oritavancin combined with ceftaroline, ampicillin, or ertapenem was evaluated for lethal synergy against VRE in 24-h time-kill assays. Oritavancin at 0.5× the MIC was combined with BLs at 0.5× the MIC or the biological free peak concentration, whichever one was lower. Synergy was defined as a ≥2-log10-CFU/ml difference between the killing achieved with the combination and that achieved with the most active single agent at 24 h. Oritavancin MICs were ≤0.125 μg/ml for all MRSA isolates except three VISA isolates with MICs of 0.25 μg/ml. Oritavancin MICs for VRE ranged from 0.03 to 0.125 μg/ml. Oritavancin in combination with ceftaroline was synergistic against all MRSA phenotypes and statistically superior to all other combinations against DNS MRSA, hVISA, and MRSA isolates (P< 0.02). Oritavancin in combination with cefazolin and oritavancin in combination with nafcillin were also synergistic against all MRSA strains. Synergy between oritavancin and all BLs was revealed against VRE strain 8019, while synergy between oritavancin and ampicillin or ertapenem but not ceftaroline was demonstrated against VRE strain R7164. The data support the potential use of oritavancin in combination

  5. Artesunate has its enhancement on antibacterial activity of β-lactams via increasing the antibiotic accumulation within methicillin-resistant Staphylococcus aureus (MRSA).

    PubMed

    Jiang, Weiwei; Li, Bin; Zheng, Xinchuan; Liu, Xin; Pan, Xichun; Qing, Rongxin; Cen, Yanyan; Zheng, Jiang; Zhou, Hong

    2013-06-01

    Methicillin-resistant Staphylococcus aureus (MRSA) has now emerged as a predominant and serious pathogen because of its resistance to a large group of antibiotics, leading to high morbidity and mortality. Therefore, to develop new agents against resistance is urgently required. Previously, artesunate (AS) was found to enhance the antibacterial effect of β-lactams against MRSA. In this study, AS was first found to increase the accumulation of antibiotics (daunorubicin and oxacillin) within MRSA by laser confocal microscopy and liquid chromatography-tandem MS method, suggesting the increased antibiotics accumulation might be related to the enhancement of AS on antibiotics. Furthermore, AS was found not to destroy the cell structure of MRSA by transmission electron microscope. AS was found to inhibit gene expressions of important efflux pumps such as NorA, NorB and NorC, but not MepA, SepA and MdeA. In conclusion, our results showed that AS was capable of enhancing the antibacterial activity of β-lactams via increasing antibiotic accumulations within MRSA through inhibiting gene expressions of efflux pumps such as NorA, NorB and NorC, but did not destroy the cell structure of MRSA. AS could be further investigated as a candidate drug for treatment of MRSA infection.

  6. Substitution of Thr for Ala-237 in TEM-17, TEM-12 and TEM-26: alterations in beta-lactam resistance conferred on Escherichia coli.

    PubMed

    Giakkoupi, P; Hujer, A M; Miriagou, V; Tzelepi, E; Bonomo, R A; Tzouvelekis, L S

    2001-07-10

    Non-naturally occurring mutants of TEM-17 (E104K), TEM-12 (R164S) and TEM-26 (E104K:R164S) extended-spectrum (ES) beta-lactamases bearing threonine at position 237 were constructed by site-specific mutagenesis and expressed under isogenic conditions in Escherichia coli. Quantification of beta-lactamase activities and immunoblotting indicated that Ala-237-->Thr did not significantly affect expression levels of these ES enzymes. Minimum inhibitory concentrations of beta-lactam antibiotics showed that the presence of threonine at position 237 exerted a dominant effect increasing the enzymes' preference for various early generation cephalosporins over penicillins. Activity against broad-spectrum oxyimino-beta-lactams was also changed. The effect of Ala-237-->Thr on the activity against ceftazidime, aztreonam, cefepime and cefpirome of all three ES TEM enzymes was detrimental. Introduction of Thr-237 improved activity against cefotaxime and ceftriaxone in TEM-12 and TEM-26, but not in TEM-17. PMID:11445164

  7. A Polysaccharide Deacetylase Gene (pdaA) Is Required for Germination and for Production of Muramic δ-Lactam Residues in the Spore Cortex of Bacillus subtilis

    PubMed Central

    Fukushima, Tatsuya; Yamamoto, Hiroki; Atrih, Abdelmadjid; Foster, Simon J.; Sekiguchi, Junichi

    2002-01-01

    The predicted amino acid sequence of Bacillus subtilis yfjS (renamed pdaA) exhibits high similarity to those of several polysaccharide deacetylases. β-Galactosidase fusion experiments and results of Northern hybridization with sporulation sigma mutants indicated that the pdaA gene is transcribed by EσG RNA polymerase. pdaA-deficient spores were bright by phase-contrast microscopy, and the spores were induced to germination on the addition of l-alanine. Germination-associated spore darkening, a slow and partial decrease in absorbance, and slightly lower dipicolinic acid release compared with that by the wild-type strain were observed. In particular, the release of hexosamine-containing materials was lacking in the pdaA mutant. Muropeptide analysis indicated that the pdaA-deficient spores completely lacked muramic δ-lactam. A pdaA-gfp fusion protein constructed in strain 168 and pdaA-deficient strains indicated that the protein is localized in B. subtilis spores. The biosynthetic pathway of muramic δ-lactam is discussed. PMID:12374835

  8. Staphylococcus pseudintermedius and Staphylococcus schleiferi Subspecies coagulans from Canine Pyoderma Cases in Grenada, West Indies, and Their Susceptibility to Beta-Lactam Drugs

    PubMed Central

    Gibson, Kathryn; Frankie, Matthew; Matthew, Vanessa; Daniels, Joshua; Martin, Nancy A.; Andrews, Linton; Paterson, Tara; Sharma, Ravindra N.

    2014-01-01

    Over a 2-year period 66 cases of canine pyoderma in Grenada, West Indies, were examined by aerobic culture in order to ascertain the bacteria involved and their antimicrobial resistance patterns. Of the 116 total bacterial isolates obtained, the majority belonged to Gram-positive species, and the most common organism identified through biochemical and molecular methods was Staphylococcus pseudintermedius. Additionally, identification of a Staphylococcus schleiferi subspecies coagulans isolate was confirmed by molecular methods. All isolates of staphylococci were susceptible to beta-lactam drugs: amoxicillin-clavulanic acid, cefovecin, cefoxitin, cefpodoxime, and cephalothin. They were also susceptible to chloramphenicol and enrofloxacin. Resistance was highest to tetracycline. Methicillin resistance was not detected in any isolate of S. pseudintermedius or in S. schleiferi. Among the Gram-negative bacteria, the most common species was Klebsiella pneumoniae, followed by Acinetobacter baumannii/calcoaceticus. The only drug to which all Gram-negative isolates were susceptible was enrofloxacin. This report is the first to confirm the presence of S. pseudintermedius and S. schleiferi subspecies coagulans, in dogs with pyoderma in Grenada, and the susceptibility of staphylococcal isolates to the majority of beta-lactam drugs used in veterinary practice. PMID:24592351

  9. Bioactivity Studies of β-Lactam Derived Polycyclic Fused Pyrroli-Dine/Pyrrolizidine Derivatives in Dentistry: In Vitro, In Vivo and In Silico Studies

    PubMed Central

    Winfred, Sofi Beaula; Mannivanan, Bhavani; Bhoopalan, Hemadev; Shankar, Venkatesh; Sekar, Sathiya; Venkatachalam, Deepa Parvathi; Pitani, Ravishankar; Nagendrababu, Venkateshbabu; Thaiman, Malini; Devivanayagam, Kandaswamy; Jayaraman, Jeyakanthan; Ragavachary, Raghunathan; Venkatraman, Ganesh

    2015-01-01

    The antibacterial activity of β-lactam derived polycyclic fused pyrrolidine/pyrrolizidine derivatives synthesized by 1, 3-dipolar cycloaddition reaction was evaluated against microbes involved in dental infection. Fifteen compounds were screened; among them compound 3 showed efficient antibacterial activity in an ex vivo dentinal tubule model and in vivo mice infectious model. In silico docking studies showed greater affinity to penicillin binding protein. Cell damage was observed under Scanning Electron Microscopy (SEM) which was further proved by Confocal Laser Scanning Microscope (CLSM) and quantified using Flow Cytometry by PI up-take. Compound 3 treated E. faecalis showed ROS generation and loss of membrane integrity was quantified by flow cytometry. Compound 3 was also found to be active against resistant E. faecalis strains isolated from failed root canal treatment cases. Further, compound 3 was found to be hemocompatible, not cytotoxic to normal mammalian NIH 3T3 cells and non mutagenic. It was concluded that β-lactam compound 3 exhibited promising antibacterial activity against E. faecalis involved in root canal infections and the mechanism of action was deciphered. The results of this research can be further implicated in the development of potent antibacterial medicaments with applications in dentistry. PMID:26185985

  10. Combined Experimental and Theoretical Study on Hydrogen-Bonded Complexes between Cyclic Ketones, Lactones, and Lactams with 3,4-Dinitrophenol

    NASA Astrophysics Data System (ADS)

    Esseffar, M.; Firdoussi, A. El; Bouab, W.; Abboud, J.-L. M.; Mó, O.; Yáñez, M.

    2009-10-01

    The interaction of 3,4 dinitrophenol (DNP) with cyclic ketones, lactones, and lactams was investigated by UV-visible spectroscopy and density functional theory (DFT) methods. Equilibrium constants KHB for 1:1 hydrogen bonded complexes were determined in solution in CCl4 and C6H12. For the entire range of studied bases, the pKHB scale, varying between 2.92 for β-propiolactone to 5.53 for 1-methyl-γ-caprolactam, indicates that the basicity increases with the ring size. Geometries, energies, and vibrational characteristics of complexes were obtained by means of DFT calculations. For lactones and lactams, the energy difference between the two most stable conformers, cis and trans, with respect to the ring oxygen (nitrogen) atom, is relatively small, suggesting that the complex observed in solution is probably an equilibrium mixture of both forms. The good correlation between Gibbs free energies in solution and in the gas phase, computed at the B3LYP/6-311++G(3df,2p) level of theory, confirms the reliability of our results. The electron density of the complexes has been analyzed by means of the atoms in molecules (AIM) theory and the natural bond orbital (NBO) method have been used to characterize the orbital interactions. Our theoretical survey shows that the 1:1 complexes are stabilized by a network of conventional and/or nonconventional intermolecular hydrogen bonds.

  11. Beta-lactam biosynthesis in a gram-negative eubacterium: purification and characterization of isopenicillin N synthase from Flavobacterium sp. strain SC 12.154.

    PubMed Central

    Palissa, H; von Döhren, H; Kleinkauf, H; Ting, H H; Baldwin, J E

    1989-01-01

    The occurrence, localization, and extraction of isopenicillin N-synthase (IPNS) were investigated in the gram-negative low-level beta-lactam producer Flavobacterium sp. strain SC 12.154, which forms deacetoxycephalosporin and excretes the cephabacin 7-formamidocephalosporin. IPNS was detected with anti-IPNS antibodies raised against the Cephalosporium acremonium enzyme. The flavobacterium enzyme, whose molecular mass (38 kilodaltons) and cofactor requirements resemble those of the fungal and Streptomyces enzymes, is formed at the transition from growth to the stationary phase. It was extracted into the polyethylene glycol phase of a polyethylene glycol-Ficoll-dextran three-phase system and was purified by quaternary aminoethyl ion-exchange chromatography, gel filtration, covalent chromatography on cystamine-Sepharose, and fast-protein liquid chromatography on Mono Q. The enzyme was characterized with respect to sulfhydryl requirement, inhibition by disulfides and metal ions, pH and temperature dependence, and stimulation by polyethylene glycol and low Triton X-100 concentrations, as well as by several amino acids, including alpha-aminoadipic acid and cysteine. The Km for alpha-aminoadipyl-cysteinyl-D-valine was 0.08 mM. An inactive membrane-associated form of IPNS was detected together with a beta-lactamase active on isopenicillin N. The system has been suggested as a model for the study of endogenous functions of beta-lactams in bacteria. Images PMID:2793834

  12. Resistance to cefotaxime and seven other beta-lactams in members of the family Enterobacteriaceae: a 3-year survey in France.

    PubMed Central

    Sirot, D L; Goldstein, F W; Soussy, C J; Courtieu, A L; Husson, M O; Lemozy, J; Meyran, M; Morel, C; Perez, R; Quentin-Noury, C

    1992-01-01

    During the second quarter each of 1988, 1989, and 1990, a French collaborative study group, including 12 university hospital laboratories, surveyed the resistance to beta-lactams of clinical isolates from hospitalized patients: consecutively, 10,641, 10,692, and 9,382 isolates were tested. The distribution of bacterial species over time was similar in each laboratory. The susceptibilities of microorganisms to amoxicillin, ticarcillin, cephalothin, cefoxitin, cefotaxime (CTX), ceftazidime (CAZ), aztreonam (ATM), and imipenem (IPM) were measured by the disk diffusion method in accordance with the recommendations of the Antibiogram Committee of the French Society for Microbiology. Five reference strains were included for quality control. Extended-spectrum beta-lactamases were detected by the synergistic effect of the combination of clavulanic acid-amoxicillin with CTX, CAZ, and ATM in the double-diffusion test. A synergistic effect with CTX, CAZ, and ATM was detected for 1.5% of all strains, mainly those of Klebsiella pneumoniae (13.3%). For this species, the synergy test enabled the detection of roughly 50% of the resistant strains misclassified as susceptible on the basis of interpretative standards. Extended-spectrum beta-lactamases disseminated in 1990 in most enterobacterial species but at a low frequency. Important variations in the percentages of resistant strains were observed in terms of bacterial species, hospitals, and wards. However, when the total number of strains was considered, the percentages of resistance to newer beta-lactams remained low. PMID:1416850

  13. Bioactivity Studies of β-Lactam Derived Polycyclic Fused Pyrroli-Dine/Pyrrolizidine Derivatives in Dentistry: In Vitro, In Vivo and In Silico Studies.

    PubMed

    Meiyazhagan, Gowri; Raju, Rajesh; Winfred, Sofi Beaula; Mannivanan, Bhavani; Bhoopalan, Hemadev; Shankar, Venkatesh; Sekar, Sathiya; Venkatachalam, Deepa Parvathi; Pitani, Ravishankar; Nagendrababu, Venkateshbabu; Thaiman, Malini; Devivanayagam, Kandaswamy; Jayaraman, Jeyakanthan; Ragavachary, Raghunathan; Venkatraman, Ganesh

    2015-01-01

    The antibacterial activity of β-lactam derived polycyclic fused pyrrolidine/pyrrolizidine derivatives synthesized by 1, 3-dipolar cycloaddition reaction was evaluated against microbes involved in dental infection. Fifteen compounds were screened; among them compound 3 showed efficient antibacterial activity in an ex vivo dentinal tubule model and in vivo mice infectious model. In silico docking studies showed greater affinity to penicillin binding protein. Cell damage was observed under Scanning Electron Microscopy (SEM) which was further proved by Confocal Laser Scanning Microscope (CLSM) and quantified using Flow Cytometry by PI up-take. Compound 3 treated E. faecalis showed ROS generation and loss of membrane integrity was quantified by flow cytometry. Compound 3 was also found to be active against resistant E. faecalis strains isolated from failed root canal treatment cases. Further, compound 3 was found to be hemocompatible, not cytotoxic to normal mammalian NIH 3T3 cells and non mutagenic. It was concluded that β-lactam compound 3 exhibited promising antibacterial activity against E. faecalis involved in root canal infections and the mechanism of action was deciphered. The results of this research can be further implicated in the development of potent antibacterial medicaments with applications in dentistry. PMID:26185985

  14. β-Lactam antibiotics. Spectroscopy and molecular orbital (MO) calculations . Part I: IR studies of complexation in penicillin-transition metal ion systems and semi-empirical PM3 calculations on simple model compounds

    NASA Astrophysics Data System (ADS)

    Kupka, Teobald

    1997-12-01

    IR studies were preformed to determine possible transition metal ion binding sites of penicillin. the observed changes in spectral position and shape of characteristic IR bands of cloxacillin in the presence of transition metal ions (both in solutions and in the solid state) indicate formation of M-L complexes with engagement of -COO - and/or -CONH- functional groups. The small shift of νCO towards higher frequencies rules out direct M-L interaction via β-lactam carbonyl. PM3 calculations on simple model compounds (substituted formamide, cyclic ketones, lactams and substituted monocyclic β-lactams) have been performed. All structures were fully optimized and the calculated bond lengths, angles, heats of formation and CO stretching frequencies were discussed to determine the β-lactam binding sites and to explain its susceptibility towards nucleophilic attack (hydrolysis in vitro) and biological activity. The relative changes of calculated values were critically compared with available experimental data and same correlation between structural parameters and in vivo activity was shown.

  15. Degradation kinetics and mechanism of β-lactam antibiotics by the activation of H2O2 and Na2S2O8 under UV-254nm irradiation.

    PubMed

    He, Xuexiang; Mezyk, Stephen P; Michael, Irene; Fatta-Kassinos, Despo; Dionysiou, Dionysios D

    2014-08-30

    The extensive production and usage of antibiotics have led to an increasing occurrence of antibiotic residuals in various aquatic compartments, presenting a significant threat to both ecosystem and human health. This study investigated the degradation of selected β-lactam antibiotics (penicillins: ampicillin, penicillin V, and piperacillin; cephalosporin: cephalothin) by UV-254nm activated H2O2 and S2O8(2-) photochemical processes. The UV irradiation alone resulted in various degrees of direct photolysis of the antibiotics; while the addition of the oxidants improved significantly the removal efficiency. The steady-state radical concentrations were estimated, revealing a non-negligible contribution of hydroxyl radicals in the UV/S2O8(2-) system. Mineralization of the β-lactams could be achieved at high UV fluence, with a slow formation of SO4(2-) and a much lower elimination of total organic carbon (TOC). The transformation mechanisms were also investigated showing the main reaction pathways of hydroxylation (+16Da) at the aromatic ring and/or the sulfur atom, hydrolysis (+18Da) at the β-lactam ring and decarboxylation (-44Da) for the three penicillins. Oxidation of amine group was also observed for ampicillin. This study suggests that UV/H2O2 and UV/S2O8(2-) advanced oxidation processes (AOPs) are capable of degrading β-lactam antibiotics decreasing consequently the antibiotic activity of treated waters.

  16. In vitro and in vivo antibacterial activities of cranberry press cake extracts alone or in combination with β-lactams against Staphylococcus aureus

    PubMed Central

    2013-01-01

    Background Cranberry fruits possess many biological activities partly due to their various phenolic compounds; however the underlying modes of action are poorly understood. We studied the effect of cranberry fruit extracts on the gene expression of Staphylococcus aureus to identify specific cellular processes involved in the antibacterial action. Methods Transcriptional profiles of four S. aureus strains grown in broth supplemented or not with 2 mg/ml of a commercial cranberry preparation (Nutricran®90) were compared using DNA arrays to reveal gene modulations serving as markers for biological activity. Ethanol extracted pressed cakes from fresh fruits also produced various fractions and their effects on marker genes were demonstrated by qPCR. Minimal inhibitory concentrations (MICs) of the most effective cranberry fraction (FC111) were determined against multiple S. aureus strains and drug interactions with β-lactam antibiotics were also evaluated. Incorporation assays with [3H]-radiolabeled precursors were performed to evaluate the effect of FC111 on DNA, RNA, peptidoglycan (PG) and protein biosynthesis. Results Treatment of S. aureus with Nutricran®90 or FC111 revealed a transcriptional signature typical of PG-acting antibiotics (up-regulation of genes vraR/S, murZ, lytM, pbp2, sgtB, fmt). The effect of FC111 on PG was confirmed by the marked inhibition of incorporation of D-[3H]alanine. The combination of β-lactams and FC111 in checkerboard assays revealed a synergistic activity against S. aureus including strain MRSA COL, which showed a 512-fold drop of amoxicillin MIC in the presence of FC111 at MIC/8. Finally, a therapeutic proof of concept was established in a mouse mastitis model of infection. S. aureus-infected mammary glands were treated with amoxicillin, FC111 or a combination of both; only the combination significantly reduced bacterial counts from infected glands (P<0.05) compared to the untreated mice. Conclusions The cranberry fraction FC111

  17. Restoration of Susceptibility of Intracellular Methicillin-Resistant Staphylococcus aureus to β-Lactams: Comparison of Strains, Cells, and Antibiotics▿ †

    PubMed Central

    Lemaire, Sandrine; Olivier, Aurélie; Van Bambeke, Françoise; Tulkens, Paul M.; Appelbaum, Peter C.; Glupczynski, Youri

    2008-01-01

    Staphylococcus aureus invades eukaryotic cells. When methicillin-resistant S. aureus (MRSA) ATCC 33591 is phagocytized by human THP-1 macrophages, complete restoration of susceptibility to cloxacillin and meropenem is shown and the strain becomes indistinguishable from MSSA ATCC 25923 due to the acid pH prevailing in phagolysosomes (S. Lemaire et al., Antimicrob. Agents Chemother. 51:1627-1632, 2007). We examined whether this observation can be extended to (i) strains of current clinical and epidemiological interest (three hospital-acquired MRSA [HA-MRSA] strains, two community-acquired MRSA [CA-MRSA] strains, two HA-MRSA strains with the vancomycin-intermediate phenotype, one HA-MRSA strain with the vancomycin-resistant phenotype, and one animal [porcine] MRSA strain), (ii) activated THP-1 cells and nonprofessional phagocytes (keratinocytes, Calu-3 bronchial epithelial cells), and (iii) other β-lactams (imipenem, oxacillin, cefuroxime, cefepime). All strains showed (i) a marked reduction in MICs in broth at pH 5.5 compared with the MIC at pH 7.4 and (ii) sigmoidal dose-response curves with cloxacillin (0.01× to 100× MIC, 24 h of incubation) after phagocytosis by THP-1 macrophages that were indistinguishable from each other and from the dose-response curve for methicillin-susceptible S. aureus (MSSA) ATCC 25923 (relative potency [50% effect], 6.09× MIC [95% confidence interval {CI}, 4.50 to 8.25]; relative efficacy [change in bacterial counts over the original inoculum for an infinitely large cloxacillin concentration, or maximal effect], −0.69 log CFU [95% CI, −0.79 to −0.58]). Similar dose-response curves for cloxacillin were also observed with MSSA ATCC 25923 and MRSA ATCC 33591 after phagocytosis by activated THP-1 macrophages, keratinocytes, and Calu-3 cells. By contrast, there was a lower level of restoration of susceptibility of MRSA ATCC 33591 to cefuroxime and cefepime after phagocytosis by THP-1 macrophages, even when the data were normalized

  18. Antibacterial compounds of Canadian honeys target bacterial cell wall inducing phenotype changes, growth inhibition and cell lysis that resemble action of β-lactam antibiotics.

    PubMed

    Brudzynski, Katrina; Sjaarda, Calvin

    2014-01-01

    Honeys show a desirable broad spectrum activity against Gram-positive and negative bacteria making antibacterial activity an intrinsic property of honey and a desirable source for new drug development. The cellular targets and underlying mechanism of action of honey antibacterial compounds remain largely unknown. To facilitate the target discovery, we employed a method of phenotypic profiling by directly comparing morphological changes in Escherichia coli induced by honeys to that of ampicillin, the cell wall-active β-lactam of known mechanism of action. Firstly, we demonstrated the purity of tested honeys from potential β-lactam contaminations using quantitative LC-ESI-MS. Exposure of log-phase E. coli to honey or ampicillin resulted in time- and concentration-dependent changes in bacterial cell shape with the appearance of filamentous phenotypes at sub-inhibitory concentrations and spheroplasts at the MBC. Cell wall destruction by both agents, clearly visible on microscopic micrographs, was accompanied by increased permeability of the lipopolysaccharide outer membrane as indicated by fluorescence-activated cell sorting (FACS). More than 90% E. coli exposed to honey or ampicillin became permeable to propidium iodide. Consistently with the FACS results, both honey-treated and ampicillin-treated E. coli cells released lipopolysaccharide endotoxins at comparable levels, which were significantly higher than controls (p<0.0001). E. coli cells transformed with the ampicillin-resistance gene (β-lactamase) remained sensitive to honey, displayed the same level of cytotoxicity, cell shape changes and endotoxin release as ampicillin-sensitive cells. As expected, β-lactamase protected the host cell from antibacterial action of ampicillin. Thus, both honey and ampicillin induced similar structural changes to the cell wall and LPS and that this ability underlies antibacterial activities of both agents. Since the cell wall is critical for cell growth and survival, honey

  19. Antibacterial compounds of Canadian honeys target bacterial cell wall inducing phenotype changes, growth inhibition and cell lysis that resemble action of β-lactam antibiotics.

    PubMed

    Brudzynski, Katrina; Sjaarda, Calvin

    2014-01-01

    Honeys show a desirable broad spectrum activity against Gram-positive and negative bacteria making antibacterial activity an intrinsic property of honey and a desirable source for new drug development. The cellular targets and underlying mechanism of action of honey antibacterial compounds remain largely unknown. To facilitate the target discovery, we employed a method of phenotypic profiling by directly comparing morphological changes in Escherichia coli induced by honeys to that of ampicillin, the cell wall-active β-lactam of known mechanism of action. Firstly, we demonstrated the purity of tested honeys from potential β-lactam contaminations using quantitative LC-ESI-MS. Exposure of log-phase E. coli to honey or ampicillin resulted in time- and concentration-dependent changes in bacterial cell shape with the appearance of filamentous phenotypes at sub-inhibitory concentrations and spheroplasts at the MBC. Cell wall destruction by both agents, clearly visible on microscopic micrographs, was accompanied by increased permeability of the lipopolysaccharide outer membrane as indicated by fluorescence-activated cell sorting (FACS). More than 90% E. coli exposed to honey or ampicillin became permeable to propidium iodide. Consistently with the FACS results, both honey-treated and ampicillin-treated E. coli cells released lipopolysaccharide endotoxins at comparable levels, which were significantly higher than controls (p<0.0001). E. coli cells transformed with the ampicillin-resistance gene (β-lactamase) remained sensitive to honey, displayed the same level of cytotoxicity, cell shape changes and endotoxin release as ampicillin-sensitive cells. As expected, β-lactamase protected the host cell from antibacterial action of ampicillin. Thus, both honey and ampicillin induced similar structural changes to the cell wall and LPS and that this ability underlies antibacterial activities of both agents. Since the cell wall is critical for cell growth and survival, honey

  20. Application of the real-time PCR method for genotypic identification of β-lactam resistance in isolates from invasive pneumococcal diseases.

    PubMed

    Chiba, Naoko; Morozumi, Miyuki; Ubukata, Kimiko

    2012-04-01

    We sought to identify genotypic resistance classes by real-time PCR in 300 Streptococcus pneumoniae isolates from invasive pneumococcal diseases. Primers and molecular beacon probes were designed for the lytA gene, 3 pbp genes, and the mefA/ermB genes. Targeted sequences of pbp1a, pbp2x, and pbp2b genes in susceptible strain R6 corresponded to those of penicillin G-nonsusceptible strains, including sites within or adjacent to conserved amino acid motifs. If amplification did not occur, the corresponding penicillin-binding protein (PBP) was considered to possess amino acid substitution(s) affecting minimal inhibitory concentrations (MICs) of β-lactam antibiotics. Real-time PCR required 90 min or less. Strains were assigned to six genotypic classes: Genotypic penicillin-susceptible S. pneumoniae (gPSSP) with 3 normal genes (22.3%); genotypic penicillin-intermediate S. pneumoniae (gPISP) (pbp2x) with an abnormal pbp2x gene (25.3%); gPISP (pbp2b) with an abnormal pbp2b gene (7.3%); gPISP (pbp1a+2x) with abnormal pbp1a+2x genes (11.3%); gPISP (pbp2x+2b) with abnormal pbp2x+2b genes (4.7%); or genotypic penicillin-resistant S. pneumoniae (gPRSP) with 3 abnormal PBP genes (29.0%). Sensitivity and specificity of real-time PCR compared with those of conventional PCR were high, 73.7-100% and 97.7-100%, respectively. As for relationships between genotype and β-lactam MICs, 90% of MICs for every resistance class were distributed within three serial dilutions for almost all antibiotics. MICs of each β-lactam antibiotic were estimated with high probability from genotypic patterns. In conclusion, determination of genotypic classes of S. pneumoniae using rapid real-time PCR is useful in selecting effective therapeutic agents for patients with pneumococcal infection.

  1. The Effect of Sub-MIC β-Lactam Antibiotic Exposure of Pseudomonas aeruginosa Strains from People with Cystic Fibrosis in a Desiccation Survival Model.

    PubMed

    Clifton, I J; Denton, M; M'zali, F; Peckham, D G

    2011-01-01

    Prior to modern typing methods, cross-infection of P. aeruginosa between people with cystic fibrosis (CF) was felt to be rare. Recently a number of studies have demonstrated the presence of clonal strains of P. aeruginosa infecting people with CF. The aim of this study was to determine whether strains of P. aeruginosa demonstrated differences in resistance to desiccation and whether preincubation in subminimum inhibitory concentrations (MICs) of β-lactam affected desiccation resistance. The experimental data were modelled to a first-order decay model and a Weibull decay model using least squares nonlinear regression. The Weibull model was the preferred model for the desiccation survival. The presence of a mucoid phenotype promoted desiccation survival. Preincubation with antibiotics did not have a consistent effect on the strains of P. aeruginosa. Meropenem reduced desiccation resistance, whereas ceftazidime had much less effect on the strains studied.

  2. Method Based on the β-Lactamase PenPC Fluorescent Labeled for β-Lactam Antibiotic Quantification in Human Plasma

    PubMed Central

    Andresen, Max; Wong, Kwok-Yin; Leung, Yun-Chung; Wong, Wai-Ting; Chan, Pak-Ho; Andresen-Vasquez, Max; Alegria, Leyla; Silva, Camila; Tapia, Pablo; Downey, Patricio; Soto, Dagoberto

    2016-01-01

    Recently, Wong et al. have successfully developed a fluorescent biosensor based on the PenPC β-lactamase which changes its intrinsic fluorescence in presence of β-lactam antibiotics (BLAs). Here, we studied systematically this correlation among the fluorescence change of the biosensor and the concentration of different BLAs aimed at developing a novel method for estimating the concentration of a wide range of BLAs. This method showed high precision and specificity and very low interference from clinically relevant samples. We were able to monitor the pharmacokinetics of meropenem in healthy volunteers as well as in an ill animal model too, indicating that the implemented method could be suitable for clinical practice. PMID:26904674

  3. Method Based on the β-Lactamase PenPC Fluorescent Labeled for β-Lactam Antibiotic Quantification in Human Plasma.

    PubMed

    Andresen, Max; Wong, Kwok-Yin; Leung, Yun-Chung; Wong, Wai-Ting; Chan, Pak-Ho; Andresen-Vasquez, Max; Alegria, Leyla; Silva, Camila; Tapia, Pablo; Downey, Patricio; Soto, Dagoberto

    2016-01-01

    Recently, Wong et al. have successfully developed a fluorescent biosensor based on the PenPC β-lactamase which changes its intrinsic fluorescence in presence of β-lactam antibiotics (BLAs). Here, we studied systematically this correlation among the fluorescence change of the biosensor and the concentration of different BLAs aimed at developing a novel method for estimating the concentration of a wide range of BLAs. This method showed high precision and specificity and very low interference from clinically relevant samples. We were able to monitor the pharmacokinetics of meropenem in healthy volunteers as well as in an ill animal model too, indicating that the implemented method could be suitable for clinical practice. PMID:26904674

  4. All detectable high-molecular-mass penicillin-binding proteins are modified in a high-level beta-lactam-resistant clinical isolate of Streptococcus mitis.

    PubMed

    Amoroso, A; Demares, D; Mollerach, M; Gutkind, G; Coyette, J

    2001-07-01

    All detectable high-molecular-mass penicillin-binding proteins (HMM PBPs) are altered in a clinical isolate of Streptococcus mitis for which the beta-lactam MICs are increased from those previously reported in our region (cefotaxime MIC, 64 microg/ml). These proteins were hardly detected at concentrations that saturate all PBPs in clinical isolates and showed, after densitometric analysis, 50-fold-lower radiotracer binding. Resistance was related to mosaic structure in all HMM PBP-coding genes, where critical region replacement was complemented not only by substitutions already reported for the closely related Streptococcus pneumoniae but also by other specific replacements that are presumably close to the active-site serine. Mosaic structure was also presumed in a pbp1a-sensitive strain used for comparison, confirming that these structures do not unambiguously imply, by themselves, detectable critical changes in the kinetic properties of these proteins.

  5. All Detectable High-Molecular-Mass Penicillin-Binding Proteins Are Modified in a High-Level β-Lactam-Resistant Clinical Isolate of Streptococcus mitis

    PubMed Central

    Amoroso, Ana; Demares, Diego; Mollerach, Marta; Gutkind, Gabriel; Coyette, Jacques

    2001-01-01

    All detectable high-molecular-mass penicillin-binding proteins (HMM PBPs) are altered in a clinical isolate of Streptococcus mitis for which the β-lactam MICs are increased from those previously reported in our region (cefotaxime MIC, 64 μg/ml). These proteins were hardly detected at concentrations that saturate all PBPs in clinical isolates and showed, after densitometric analysis, 50-fold-lower radiotracer binding. Resistance was related to mosaic structure in all HMM PBP-coding genes, where critical region replacement was complemented not only by substitutions already reported for the closely related Streptococcus pneumoniae but also by other specific replacements that are presumably close to the active-site serine. Mosaic structure was also presumed in a pbp1a-sensitive strain used for comparison, confirming that these structures do not unambiguously imply, by themselves, detectable critical changes in the kinetic properties of these proteins. PMID:11408226

  6. Synthesis of alpha-acyl-functionalized azacycles by Pd-catalyzed cross-coupling reactions of alpha-alkoxyboronates with lactam-derived vinyl triflates.

    PubMed

    Occhiato, Ernesto G; Prandi, Cristina; Ferrali, Alessandro; Guarna, Antonio; Deagostino, Annamaria; Venturello, Paolo

    2002-10-01

    Alkoxydienyl- and alkoxystyrylboronates were used for Pd-catalyzed cross-coupling reactions with lactam-derived vinyl triflates. The hydrolysis of the coupling products with alkoxystyrylboronates provided the corresponding alpha-acyl-substituted 3,4-dihydro-(2H)-pyridines and 2,3,4,5-tetrahydroazepines in good to high yields. The hydrolysis of the coupling products with alkoxydienylboronates, performed in the presence of Amberlyst 15, resulted in a Nazarov-type cyclization that afforded hexahydro[1]pyrindin-7-ones and 3,4,5,6,7,8-hexahydro-(2H)-cyclopenta[b]azepin-8-ones. This methodology represents a novel and efficient procedure for the preparation of these classes of azacyclic compounds. PMID:12354013

  7. The ABC transporter AnrAB contributes to the innate resistance of Listeria monocytogenes to nisin, bacitracin, and various beta-lactam antibiotics.

    PubMed

    Collins, Barry; Curtis, Nicola; Cotter, Paul D; Hill, Colin; Ross, R Paul

    2010-10-01

    A mariner transposon bank was used to identify loci that contribute to the innate resistance of Listeria monocytogenes to the lantibiotic nisin. In addition to highlighting the importance of a number of loci previously associated with nisin resistance (mprF, virRS, and telA), a nisin-sensitive phenotype was associated with the disruption of anrB (lmo2115), a gene encoding the permease component of an ABC transporter. The contribution of anrB to nisin resistance was confirmed by the creation of nonpolar deletion mutants. The loss of this putative multidrug resistance transporter also greatly enhanced sensitivity to bacitracin, gallidermin, and a selection of β-lactam antibiotics. A comparison of the relative antimicrobial sensitivities of a number of mutants established the ΔanrB strain as being one of the most bacitracin-sensitive L. monocytogenes strains identified to date. PMID:20643901

  8. Rational design, synthesis, and biological evaluation of lactam-bridged gramicidin A analogues: discovery of a low-hemolytic antibacterial peptide.

    PubMed

    Mao, Ji; Kuranaga, Takefumi; Hamamoto, Hiroshi; Sekimizu, Kazuhisa; Inoue, Masayuki

    2015-03-01

    A linear peptide, gramicidin A (GA), folds into a β(6.3) -helix, functions as an ion channel in the cell membrane, and exerts antibacterial activity. Herein we describe the rational design, synthesis, and biological evaluation of lactam-bridged GA analogues. The GA analogue with a 27-membered macrolactam was found to adopt a stable β(6.3) -helical conformation and exhibits higher ion-exchange activity than GA. Furthermore, this GA analogue retains the potent antibiotic activity of GA, but its hemolytic activity and toxicity toward mammalian cells are significantly lower than those of GA. This study thus dissociates the antibacterial and hemolytic/cytotoxic activities of GA, and charts a rational path forward for the development of new ion-channel-based antibiotics.

  9. Antibacterial and synergic effects of gallic acid-grafted-chitosan with β-lactams against methicillin-resistant Staphylococcus aureus (MRSA).

    PubMed

    Lee, Dae-Sung; Eom, Sung-Hwan; Kim, Young-Mog; Kim, Hye Seon; Yim, Mi-Jin; Lee, Sang-Hoon; Kim, Do-Hyung; Je, Jae-Young

    2014-10-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is spreading worldwide, emphasizing the need to search for new antibiotics. The anti-MRSA activities of gallic acid-grafted-chitosans (GA-g-chitosans) were investigated against 2 MRSA standards and 10 MRSA clinical isolates by determining the minimum inhibitory concentrations (MICs). GA-g-chitosan (I), which has the highest gallic acid content, exhibited the strongest anti-MRSA activities, with MICs of 32-64 μg/mL. A time-kill investigation revealed that GA-g-chitosan (I) exhibited a bactericidal effect at twice the MIC, also demonstrating good thermal and pH stability. Investigation of cell envelope integrity showed the release of intracellular components with an increasing absorbance value at 260 nm, indicating cell envelope damage caused by the GA-g-chitosan (I), which was further confirmed by transmission electron microscopy. When GA-g-chitosans were combined with β-lactams, including ampicillin and penicillin, synergistic effects were observed on the 2 standard MRSA strains and on the 10 clinical isolates, with fractional inhibitory indices ranging from 0.125 to 0.625. In the time-kill dynamic confirmation test, synergistic bactericidal effects were observed for the combinations of GA-g-chitosans with β-lactams, and over 4.0 log CFU/mL reductions were observed after 24 h when combination treatment was used. These results may prove GA-g-chitosans to be a potent agent when combined with ampicillin and penicillin for the elimination of MRSA.

  10. The β-lactam resistance protein Blr, a small membrane polypeptide, is a component of the Escherichia coli cell division machinery.

    PubMed

    Karimova, Gouzel; Davi, Marilyne; Ladant, Daniel

    2012-10-01

    In Escherichia coli, cell division is performed by a multimolecular machinery called the divisome, made of 10 essential proteins and more than 20 accessory proteins. Through a bacterial two-hybrid library screen, we identified the E. coli β-lactam resistance protein Blr, a short membrane polypeptide of 41 residues, as an interacting partner of the essential cell division protein FtsL. In addition to FtsL, Blr was found to associate with several other divisomal proteins, including FtsI, FtsK, FtsN, FtsQ, FtsW, and YmgF. Using fluorescently tagged Blr, we showed that this peptide localizes to the division septum and that its colocalization requires the presence of the late division protein FtsN. Although Blr is not essential, previous studies have shown that the inactivation of the blr gene increased the sensitivity of bacteria to β-lactam antibiotics or their resistance to cell envelope stress. Here, we found that Blr, when overproduced, restores the viability of E. coli ftsQ1(Ts) cells, carrying a thermosensitive allele of the ftsQ gene, during growth under low-osmotic-strength conditions (e.g., in synthetic media or in Luria-Bertani broth without NaCl). In contrast, the inactivation of blr increases the osmosensitivity of ftsQ1(Ts) cells, and blr ftsQ1 double mutants exhibit filamentous growth in LB broth even at a moderate salt concentration (0.5% NaCl) compared to parental ftsQ1(Ts) cells. Altogether, our results suggest that the small membrane polypeptide Blr is a novel component of the E. coli cell division apparatus involved in the stabilization of the divisome under certain stress conditions.

  11. Molecular evolution of beta-lactam-resistant Haemophilus influenzae: 9-year surveillance of penicillin-binding protein 3 mutations in isolates from Japan.

    PubMed

    Sanbongi, Yumiko; Suzuki, Takahisa; Osaki, Yumi; Senju, Nami; Ida, Takashi; Ubukata, Kimiko

    2006-07-01

    A total of 621 clinical isolates of Haemophilus influenzae collected in Japan between 1995 and 2003 were studied for their susceptibilities to several antimicrobial agents, beta-lactamase production, and amino acid substitutions in penicillin-binding protein 3 (PBP 3). Over the four study periods (first period, 1995 to 1996; second period, 1997 to 1998; third period, 2000 to 2001; fourth period, 2002 to 2003), the susceptibilities to beta-lactam agents decreased and the incidence of isolates with substitutions at positions 377, 385, 389, 517, and/or 526 in PBP 3 increased from 28.8% to 52.0%. Five hundred seventy-one beta-lactamase-nonproducing isolates were grouped into 18 classes, based on the pattern of the five mutations in PBP 3. The Asp526Lys substitution led to 6.0-, 4.3-, 2.4-, and 5.4-fold increases in amoxicillin-clavulanic acid, cefdinir, cefditoren, and faropenem resistance, respectively. PBP 3 with multiple substitutions (Met377Ile, Ser385Thr, and/or Leu389Phe) together with Asp526Lys resulted in increased resistance compared to that for PBP 3 with the Asp526Lys substitution alone. These results indicate that mutations at these five positions increased resistance to most beta-lactams. Although a significant change in the prevalence of beta-lactamase-producing strains was not observed, the proportions of those possessing both PBP 3 alterations and beta-lactamase production have slightly increased (from 1.4% to 5.0%). The ROB-1 beta-lactamase was rare, but this is the first report of this beta-lactamase in Japan. PMID:16801430

  12. Whole-genome sequencing reveals a link between β-lactam resistance and synthetases of the alarmone (p)ppGpp in Staphylococcus aureus.

    PubMed

    Mwangi, Michael M; Kim, Choonkeun; Chung, Marilyn; Tsai, Jennifer; Vijayadamodar, Govindan; Benitez, Michelle; Jarvie, Thomas P; Du, Lei; Tomasz, Alexander

    2013-06-01

    The overwhelming majority of methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates exhibit a peculiar heterogeneous resistance to β-lactam antibiotics: in cultures of such strains, the majority of cells display only a low level of methicillin resistance--often close to the MIC breakpoint of susceptible strains. Yet, in the same cultures, subpopulations of bacteria exhibiting very high levels of resistance are also present with variable frequencies, which are characteristic of the particular MRSA lineage. The mechanism of heterogeneous resistance is not understood. We describe here an experimental system for exploring the mechanism of heterogeneous resistance. Copies of the resistance gene mecA cloned into a temperature-sensitive plasmid were introduced into the fully sequenced methicillin-susceptible clinical isolate S. aureus strain 476. Transductants of strain 476 expressed methicillin resistance in a heterogeneous fashion: the great majority of cells showed only low MIC (0.75 μg/ml) for the antibiotic, but a minority population of highly resistant bacteria (MIC >300 μg/ml) was also present with a frequency of ∼10(-4). The genetic backgrounds of the majority and minority cells were compared by whole-genome sequencing: the only differences detectable were two point mutations in relA of the highly resistant minority population of bacteria. The relA gene codes for the synthesis of (p)ppGpp, an effector of the stringent stress response. Titration of (p)ppGpp showed increased amounts of this effector in the highly resistant cells. Involvement of (p)ppGpp synthesis genes may explain some of the perplexing aspects of β-lactam resistance in MRSA, since many environmental and genetic changes can modulate cellular levels of (p)ppGpp.

  13. A vacuolar membrane protein affects drastically the biosynthesis of the ACV tripeptide and the beta-lactam pathway of Penicillium chrysogenum.

    PubMed

    Fernández-Aguado, Marta; Teijeira, Fernando; Martín, Juan F; Ullán, Ricardo V

    2013-01-01

    The knowledge about enzymes' compartmentalization and transport processes involved in the penicillin biosynthesis in Penicillium chrysogenum is very limited. The genome of this fungus contains multiple genes encoding transporter proteins, but very little is known about them. A bioinformatic search was made to find major facilitator supefamily (MFS) membrane proteins related to CefP transporter protein involved in the entry of isopenicillin N to the peroxisome in Acremonium chrysogenum. No strict homologue of CefP was observed in P. chrysogenum, but the penV gene was found to encode a membrane protein that contained 10 clear transmembrane spanners and two other motifs COG5594 and DUF221, typical of membrane proteins. RNAi-mediated silencing of penV gene provoked a drastic reduction of the production of the δ-(L-α-aminoadipyl-L-cysteinyl-D-valine) (ACV) and isopenicillin N intermediates and the final product of the pathway. RT-PCR and northern blot analyses confirmed a reduction in the expression levels of the pcbC and penDE biosynthetic genes, whereas that of the pcbAB gene increased. Localization studies by fluorescent laser scanning microscopy using Dsred and GFP fluorescent fusion proteins and the FM 4-64 fluorescent dye showed clearly that the protein was located in the vacuolar membrane. These results indicate that PenV participates in the first stage of the beta-lactam biosynthesis (i.e., the formation of the ACV tripeptide), probably taking part in the supply of amino acids from the vacuolar lumen to the vacuole-anchored ACV synthetase. This is in agreement with several reports on the localization of the ACV synthetase and provides increased evidence for a compartmentalized storage of precursor amino acids for non-ribosomal peptides. PenV is the first MFS transporter of P. chrysogenum linked to the beta-lactam biosynthesis that has been located in the vacuolar membrane.

  14. Effect of clavulanic acid on activity of beta-lactam antibiotics in Serratia marcescens isolates producing both a TEM beta-lactamase and a chromosomal cephalosporinase.

    PubMed Central

    Bush, K; Flamm, R K; Ohringer, S; Singer, S B; Summerill, R; Bonner, D P

    1991-01-01

    An isolate of Serratia marcescens that produced both an inducible chromosomal and a plasmid-mediated TEM-1 beta-lactamase was resistant to ampicillin and amoxicillin and also demonstrated decreased susceptibility to extended-spectrum beta-lactam antibiotics (ESBAs). Clavulanic acid did not lower the MICs of the ESBAs, but it decreased the MICs of the penicillins. The TEM-1-producing plasmid was transferred to a more susceptible S. marcescens strain that produced a well-characterized inducible chromosomal beta-lactamase. The MICs of the ESBAs remained at a low level for the transconjugant. Ampicillin and amoxicillin which were good substrates for the plasmid-mediated enzyme, were not well hydrolyzed by the chromosomal enzymes; the ESBAs were hydrolyzed slowly by all the enzymes. When each of the S. marcescens strains was grown with these beta-lactam antibiotics, at least modest increases in chromosomal beta-lactamase activity were observed. When organisms were grown in the presence of clavulanic acid and an ESBA, no enhanced induction was observed. The increases in the MICs of the ESBAs observed for the initial clinical isolate may have been due to a combination of low inducibility, slow hydrolysis, and differences in permeability between the S. marcescens isolates. When clavulanic acid and a penicillin were added to strains that produced both a plasmid-mediated TEM and a chromosomal beta-lactamase, much higher levels of chromosomal beta-lactamase activity were present than were observed in cultures induced by the penicillin alone. This was due to the higher levels of penicillin that were available for induction as a result of inhibition of the TEM enzyme by clavulanate. Images PMID:1803992

  15. Divergent, stereoselective access to heterocyclic α,α-quaternary- and β(2,3,3)-amino acid derivatives from a N-Pmp-protected Orn-derived β-lactam.

    PubMed

    Núñez-Villanueva, Diego; García-López, M Teresa; Martín-Martínez, Mercedes; González-Muñiz, Rosario

    2015-05-14

    A suitably protected Orn-derived (3S,4S)-β-lactam was used as common intermediate in the synthesis of conformationally constrained (3S,4S)-2-oxoazepane α,α- and (2S,3S)-2-oxopiperidine-β(2,3,3)-amino acid derivatives. Compared to alternative procedures using an N-p-methoxybenzyl group at the 2-azetidinone, the incorporation of a p-methoxyphenyl moiety is crucial for the excellent stereochemical outcomes in the preparation of these heterocyclic amino acids. Chemoselective 7- or 6-exo-trig cyclization was achieved through alternative sequences of Pmp-deprotection/Boc-activation, followed by inter- and intramolecular β-lactam ring opening, respectively.

  16. Biochemical characterization of penicillin-resistant and -sensitive penicillin-binding protein 2x transpeptidase activities of Streptococcus pneumoniae and mechanistic implications in bacterial resistance to beta-lactam antibiotics.

    PubMed Central

    Zhao, G; Yeh, W K; Carnahan, R H; Flokowitsch, J; Meier, T I; Alborn, W E; Becker, G W; Jaskunas, S R

    1997-01-01

    To understand the biochemical basis of resistance of bacteria to beta-lactam antibiotics, we purified a penicillin-resistant penicillin-binding protein 2x (R-PBP2x) and a penicillin-sensitive PBP2x (S-PBP2x) enzyme of Streptococcus pneumoniae and characterized their transpeptidase activities, using a thioester analog of stem peptides as a substrate. A comparison of the k(cat)/Km values for the two purified enzymes (3,400 M(-1) s(-1) for S-PBP2x and 11.2 M(-1) s(-1) for R-PBP2x) suggests that they are significantly different kinetically. Implications of this finding are discussed. We also found that the two purified enzymes did not possess a detectable level of beta-lactam hydrolytic activity. Finally, we show that the expression levels of both PBP2x enzymes were similar during different growth phases. PMID:9244281

  17. One-Pot Conversion of N-Allyl-α-cyano Esters to α-Allyl-α-cyano Lactams through a Hydrolysis/Ketene Formation/Cyclization/Claisen Rearrangement Sequence.

    PubMed

    Shen, Mei-Hua; Han, Mei; Xu, Hua-Dong

    2016-03-01

    An intramolecular ketene aza-Claisen rearrangement is developed for the first time to enable the stereoselective synthesis of α-ally-α-cyano-lactams from N-allyl amino esters. This reaction also exhibits outstanding chemoselectivity when an unsymmetrical bis-N-allyl group is present in the starting molecule. The usefulness of this method is demonstrated by a short synthesis of optically active bicyclolactam from l-proline. PMID:26872217

  18. Validation of the charm 3 SL3 beta-lactam test for screening raw milk in compliance with the U.S. pasteurized milk ordinance. Performance Tested Method 071002.

    PubMed

    Salter, Robert S; Douglas, David; McRobbie, Lindsey; Quintana, Julio; Legg, David; Schwartz, Janine; Conaway, David; McPhee, Carla; Saul, Steven; Markovsky, Robert

    2011-01-01

    The Charm 3 SL3 beta-Lactam Test is a 3 min receptor-based lateral-flow Rapid One-Step Assay (ROSA) that detects the six beta-lactam drugs of concern approved for dairy cattle in the United States. The method is a biochemical formulation change of the SL3 beta-Lactam Test evaluated and approved in 2007. The Charm 3 SL3 was evaluated under the AOAC Research Institute Performance Tested Method (PTM) program following the protocol of the U.S. Food and Drug Administration, Center for Veterinary Medicine. The method was approved as PTM 071002 on May 8, 2009. The following drugs were detected in three combined lots: penicillin G at 3.8 ppb, ampicillin at 8.0 ppb, amoxicillin at 8.4 ppb, cephapirin at 20.0 ppb, ceftiofur (total metabolites) at 79 ppb, and cloxacillin at 8.6 ppb > or = 90% of the time with 95% confidence. These detection levels are lower than, but within 75% of, the U.S. Safe Level/Tolerances. Lot-to-lot repeatability was typically within 20% of these determined levels. The test kit was found to be suitable for testing thawed frozen samples. It was also found to respond with equal or better sensitivity to samples that contained incurred analytes, i.e., both the microbiologically active parent drug and its active metabolites. There were no interferences from somatic cells at 1.1 million/mL, bacterial cells at 300 000 CFU/mL, or 32 other non-beta-lactam drugs at 100 ppb. Ruggedness experiments indicated that the test procedure is robust. These results meet the fit-for-purpose approval criteria for inclusion in the National Conference for Interstate Milk Shipments milk testing program.

  19. Novel plasmid-mediated beta-lactamase (MIR-1) conferring resistance to oxyimino- and alpha-methoxy beta-lactams in clinical isolates of Klebsiella pneumoniae.

    PubMed

    Papanicolaou, G A; Medeiros, A A; Jacoby, G A

    1990-11-01

    Klebsiella pneumoniae isolates from 11 patients at the Miriam Hospital were identified as resistant to cefoxitin and ceftibuten as well as to aztreonam, cefotaxime, and ceftazidime. Resistance could be transferred by conjugation or transformation with plasmid DNA into Escherichia coli and was due to the production of a beta-lactamase with an isoelectric point of 8.4 named MIR-1. In E. coli, MIR-1 conferred resistance to aztreonam, cefotaxime, ceftazidime, ceftibuten, ceftriaxone, and such alpha-methoxy beta-lactams as cefmetazole, cefotetan, cefoxitin, and moxalactam. In vitro, MIR-1 hydrolyzed cephalothin and cephaloridine much more rapidly than it did penicillin G, ampicillin, or carbenicillin. Cefotaxime was hydrolyzed at 10% the rate of cephaloridine. Cefoxitin inactivation could only be detected by a microbiological test. The inhibition profile of MIR-1 was similar to that of chromosomally mediated class I beta-lactamases. Potassium clavulanate had little effect on cefoxitin or cefibuten resistance and was a poor inhibitor of MIR-1 activity. Cefoxitin or imipenem did not induce MIR-1. The gene determining MIR-1 was cloned on a 1.4-kb AccI-PstI fragment. Under stringent conditions, probes for TEM-1 and SHV-1 genes and the E. coli ampC gene failed to hybridize with the MIR-1 gene. However, a provisional sequence of 150 bp of the MIR-1 gene proved to be 90% identical to the sequence of ampC from Enterobacter cloacae but only 71% identical to that of E. coli, thus explaining the lack of hybridization to the E. coli ampC probe. Plasmid profiles of the 11 K. pneumoniae clinical isolates were not identical, but each contained a plasmid from 40 to 60 kb that hybridized with the cloned MIR-1 gene. Both transfer-proficient and transfer-deficient MIR-1 plasmids belonged to the N incompatibility group. Thus, the resistance of these K. pneumoniae strains was the result of plasmid acquisition of a class I beta-lactamase, a new resistance determinant that expands the kinds

  20. Bulgecin A as a β-lactam enhancer for carbapenem-resistant Pseudomonas aeruginosa and carbapenem-resistant Acinetobacter baumannii clinical isolates containing various resistance mechanisms

    PubMed Central

    Skalweit, Marion J; Li, Mei

    2016-01-01

    Genetic screening of Pseudomonas aeruginosa (PSDA) and Acinetobacter baumannii (ACB) reveals genes that confer increased susceptibility to β-lactams when disrupted, suggesting novel drug targets. One such target is lytic transglycosylase. Bulgecin A (BlgA) is a natural product of Pseudomonas mesoacidophila and a lytic transglycosolase inhibitor that works synergistically with β-lactams targeting PBP3 for Enterobacteriaceae. BlgA also weakly inhibits di-Zn2+ metallo-β-lactamases like L1 of Stenotrophomonas maltophilia. We hypothesized that because of its unique mechanism of action, BlgA could restore susceptibility to carbapenems in carbapenem-resistant PSDA (CR-PSDA) and carbapenem-resistant ACB, as well as ACB resistant to sulbactam. A BlgA-containing extract was prepared using a previously published protocol. CR-PSDA clinical isolates demonstrating a variety of carbapenem resistance mechanisms (VIM-2 carbapenemases, efflux mechanisms, and AmpC producer expression) were characterized with agar dilution minimum inhibitory concentration (MIC) testing and polymerase chain reaction. Growth curves using these strains were prepared using meropenem, BlgA extract, and meropenem plus BlgA extract. A concentrated Blg A extract combined with low concentrations of meropenem, was able to inhibit the growth of clinical strains of CR-PSDA for strains that had meropenem MICs ≥8 mg/L by agar dilution, and a clinical strain of an OXA-24 producing ACB that had a meropenem MIC >32 mg/L and intermediate ampicillin/sulbactam susceptibility. Similar experiments were conducted on a TEM-1 producing ACB strain resistant to sulbactam. BlgA with ampicillin/sulbactam inhibited the growth of this organism. As in Enterobacteriaceae, BlgA appears to restore the efficacy of meropenem in suppressing the growth of CR-PSDA and carbapenem-resistant ACB strains with a variety of common carbapenem resistance mechanisms. BlgA extract also inhibits VIM-2 β-lactamase in vitro. BlgA may prove to be

  1. Novel plasmid-mediated beta-lactamase (MIR-1) conferring resistance to oxyimino- and alpha-methoxy beta-lactams in clinical isolates of Klebsiella pneumoniae.

    PubMed Central

    Papanicolaou, G A; Medeiros, A A; Jacoby, G A

    1990-01-01

    Klebsiella pneumoniae isolates from 11 patients at the Miriam Hospital were identified as resistant to cefoxitin and ceftibuten as well as to aztreonam, cefotaxime, and ceftazidime. Resistance could be transferred by conjugation or transformation with plasmid DNA into Escherichia coli and was due to the production of a beta-lactamase with an isoelectric point of 8.4 named MIR-1. In E. coli, MIR-1 conferred resistance to aztreonam, cefotaxime, ceftazidime, ceftibuten, ceftriaxone, and such alpha-methoxy beta-lactams as cefmetazole, cefotetan, cefoxitin, and moxalactam. In vitro, MIR-1 hydrolyzed cephalothin and cephaloridine much more rapidly than it did penicillin G, ampicillin, or carbenicillin. Cefotaxime was hydrolyzed at 10% the rate of cephaloridine. Cefoxitin inactivation could only be detected by a microbiological test. The inhibition profile of MIR-1 was similar to that of chromosomally mediated class I beta-lactamases. Potassium clavulanate had little effect on cefoxitin or cefibuten resistance and was a poor inhibitor of MIR-1 activity. Cefoxitin or imipenem did not induce MIR-1. The gene determining MIR-1 was cloned on a 1.4-kb AccI-PstI fragment. Under stringent conditions, probes for TEM-1 and SHV-1 genes and the E. coli ampC gene failed to hybridize with the MIR-1 gene. However, a provisional sequence of 150 bp of the MIR-1 gene proved to be 90% identical to the sequence of ampC from Enterobacter cloacae but only 71% identical to that of E. coli, thus explaining the lack of hybridization to the E. coli ampC probe. Plasmid profiles of the 11 K. pneumoniae clinical isolates were not identical, but each contained a plasmid from 40 to 60 kb that hybridized with the cloned MIR-1 gene. Both transfer-proficient and transfer-deficient MIR-1 plasmids belonged to the N incompatibility group. Thus, the resistance of these K. pneumoniae strains was the result of plasmid acquisition of a class I beta-lactamase, a new resistance determinant that expands the kinds

  2. Ultrastructural Changes in Clinical and Microbiota Isolates of Klebsiella pneumoniae Carriers of Genes bla SHV, bla TEM, bla CTX-M, or bla KPC When Subject to β-Lactam Antibiotics.

    PubMed

    Veras, Dyana Leal; Lopes, Ana Catarina de Souza; da Silva, Grasielle Vaz; Gonçalves, Gabriel Gazzoni Araújo; de Freitas, Catarina Fernandes; de Lima, Fernanda Cristina Gomes; Maciel, Maria Amélia Vieira; Feitosa, Ana Paula Sampaio; Alves, Luiz Carlos; Brayner, Fábio André

    2015-01-01

    The aim of this study was to characterize the ultrastructural effects caused by β-lactam antibiotics in Klebsiella pneumoniae isolates. Three K. pneumoniae clinical isolates were selected for the study with resistance profiles for third-generation cephalosporins, aztreonam, and/or imipenem and with different resistance genes for extended-spectrum β-lactamases (ESBL) or Klebsiella pneumoniae carbapenemase (KPC). Two K. pneumoniae isolates obtained from the microbiota, which were both resistant to amoxicillin and ampicillin, were also analyzed. In accordance with the susceptibility profile, the clinical isolates were subjected to subminimum inhibitory concentrations (sub-MICs) of cefotaxime, ceftazidime, aztreonam, and imipenem and the isolates from the microbiota to ampicillin and amoxicillin, for analysis by means of scanning and transmission electron microscopy. The K. pneumoniae isolates showed different morphological and ultrastructural changes after subjection to β-lactams tested at different concentrations, such as cell filamentation, loss of cytoplasmic material, and deformation of dividing septa. Our results demonstrate that K. pneumoniae isolates harboring different genes that encode for β-lactamases show cell alterations when subjected to different β-lactam antibiotics, thus suggesting that they possess residual activity in vitro, despite the phenotypic resistance presented in the isolates analyzed. PMID:26491715

  3. Development and validation of a fast and uniform approach to quantify β-lactam antibiotics in human plasma by solid phase extraction-liquid chromatography-electrospray-tandem mass spectrometry.

    PubMed

    Colin, Pieter; De Bock, Lies; T'jollyn, Huybrecht; Boussery, Koen; Van Bocxlaer, Jan

    2013-01-15

    Monitoring of plasma antibiotic concentrations is necessary for individualization of antimicrobial chemotherapy dosing in special patient populations. One of these special populations of interest are the post-bariatric surgery patients. Until today, little is known on the effect of this procedure on drug disposition and efficacy. Therefore, close monitoring of antimicrobial plasma concentrations in these patients is warranted. A fast and uniform ultra-high-performance liquid chromatography (UPLC) method with tandem mass spectrometric detection (MS/MS) has been developed and qualified for the simultaneous quantification of β-lactam antibiotics in human plasma. Compounds included in this multi-component analysis are: amoxicillin, ampicillin, phenoxymethylpenicillin, piperacillin, cefuroxime, cefadroxil, flucloxacillin, meropenem, cefepime, ceftazidime, tazobactam, linezolid and cefazolin. After spiking of five different stable isotope labelled internal standards, plasma samples were prepared for UPLC-MS/MS analysis by mixed-mode solid phase extraction. The developed method was proven to be free of (relative) matrix effects and proved to be reliable for the quantification of 12 out of 13 β-lactam antibiotics. As a proof of concept the method has been applied to plasma samples obtained from a healthy volunteer treated with amoxicillin. The analytical method is suitable for use in a therapeutic drug monitoring setting, providing the clinician with reliable measurements on β-lactam antibiotic plasma concentrations in a timely manner. PMID:23200389

  4. Development and validation of a fast and uniform approach to quantify β-lactam antibiotics in human plasma by solid phase extraction-liquid chromatography-electrospray-tandem mass spectrometry.

    PubMed

    Colin, Pieter; De Bock, Lies; T'jollyn, Huybrecht; Boussery, Koen; Van Bocxlaer, Jan

    2013-01-15

    Monitoring of plasma antibiotic concentrations is necessary for individualization of antimicrobial chemotherapy dosing in special patient populations. One of these special populations of interest are the post-bariatric surgery patients. Until today, little is known on the effect of this procedure on drug disposition and efficacy. Therefore, close monitoring of antimicrobial plasma concentrations in these patients is warranted. A fast and uniform ultra-high-performance liquid chromatography (UPLC) method with tandem mass spectrometric detection (MS/MS) has been developed and qualified for the simultaneous quantification of β-lactam antibiotics in human plasma. Compounds included in this multi-component analysis are: amoxicillin, ampicillin, phenoxymethylpenicillin, piperacillin, cefuroxime, cefadroxil, flucloxacillin, meropenem, cefepime, ceftazidime, tazobactam, linezolid and cefazolin. After spiking of five different stable isotope labelled internal standards, plasma samples were prepared for UPLC-MS/MS analysis by mixed-mode solid phase extraction. The developed method was proven to be free of (relative) matrix effects and proved to be reliable for the quantification of 12 out of 13 β-lactam antibiotics. As a proof of concept the method has been applied to plasma samples obtained from a healthy volunteer treated with amoxicillin. The analytical method is suitable for use in a therapeutic drug monitoring setting, providing the clinician with reliable measurements on β-lactam antibiotic plasma concentrations in a timely manner.

  5. Ultrastructural Changes in Clinical and Microbiota Isolates of Klebsiella pneumoniae Carriers of Genes blaSHV, blaTEM, blaCTX-M, or blaKPC When Subject to β-Lactam Antibiotics

    PubMed Central

    Veras, Dyana Leal; de Souza Lopes, Ana Catarina; Vaz da Silva, Grasielle; Araújo Gonçalves, Gabriel Gazzoni; de Freitas, Catarina Fernandes; de Lima, Fernanda Cristina Gomes; Vieira Maciel, Maria Amélia; Feitosa, Ana Paula Sampaio; Alves, Luiz Carlos; Brayner, Fábio André

    2015-01-01

    The aim of this study was to characterize the ultrastructural effects caused by β-lactam antibiotics in Klebsiella pneumoniae isolates. Three K. pneumoniae clinical isolates were selected for the study with resistance profiles for third-generation cephalosporins, aztreonam, and/or imipenem and with different resistance genes for extended-spectrum β-lactamases (ESBL) or Klebsiella pneumoniae carbapenemase (KPC). Two K. pneumoniae isolates obtained from the microbiota, which were both resistant to amoxicillin and ampicillin, were also analyzed. In accordance with the susceptibility profile, the clinical isolates were subjected to subminimum inhibitory concentrations (sub-MICs) of cefotaxime, ceftazidime, aztreonam, and imipenem and the isolates from the microbiota to ampicillin and amoxicillin, for analysis by means of scanning and transmission electron microscopy. The K. pneumoniae isolates showed different morphological and ultrastructural changes after subjection to β-lactams tested at different concentrations, such as cell filamentation, loss of cytoplasmic material, and deformation of dividing septa. Our results demonstrate that K. pneumoniae isolates harboring different genes that encode for β-lactamases show cell alterations when subjected to different β-lactam antibiotics, thus suggesting that they possess residual activity in vitro, despite the phenotypic resistance presented in the isolates analyzed. PMID:26491715

  6. β-lactam antibiotics residues analysis in bovine milk by LC-ESI-MS/MS: a simple and fast liquid-liquid extraction method.

    PubMed

    Jank, L; Hoff, R B; Tarouco, P C; Barreto, F; Pizzolato, T M

    2012-01-01

    This study presents the development and validation of a simple method for the detection and quantification of six β-lactam antibiotics residues (ceftiofur, penicillin G, penicillin V, oxacillin, cloxacillin and dicloxacillin) in bovine milk using a fast liquid-liquid extraction (LLE) for sample preparation, followed by liquid chromatography-electrospray-tandem mass spectrometry (LC-MS/MS). LLE consisted of the addition of acetonitrile to the sample, followed by addition of sodium chloride, centrifugation and direct injection of an aliquot into the LC-MS/MS system. Separation was performed in a C(18) column, using acetonitrile and water, both with 0.1% of formic acid, as mobile phase. Method validation was performed according to the criteria of Commission Decision 2002/657/EC. Limits of detection ranged from 0.4 (penicillin G and penicillin V) to 10.0 ng ml(-1) (ceftiofur), and linearity was achieved. The decision limit (CCα), detection capability (CCβ), accuracy, inter- and intra-day repeatability of the method are reported.

  7. Ceftolozane/tazobactam and ceftazidime/avibactam: two novel β-lactam/β-lactamase inhibitor combination agents for the treatment of resistant Gram-negative bacterial infections.

    PubMed

    Liscio, Jordan L; Mahoney, Monica V; Hirsch, Elizabeth B

    2015-09-01

    The rise in resistant Gram-negative bacteria is a major concern and has led to difficulty in treating multidrug-resistant (MDR) infections. Two recently approved combination antibiotics, ceftolozane/tazobactam and ceftazidime/avibactam, may be effective in treating these resistant infections. Ceftolozane is a novel cephalosporin that has been developed in combination with tazobactam, a recognised β-lactamase inhibitor (BLI). Avibactam is a novel BLI combined with ceftazidime, a cephalosporin with an established history. Both of these β-lactam/BLI combination agents have been shown to retain in vitro activity against selected resistant Gram-negative pathogens, including Enterobacteriaceae and Pseudomonas aeruginosa; notably, ceftazidime/avibactam has demonstrated consistent activity against Klebsiella pneumoniae carbapenemase (KPC)-producing organisms. Both agents have been approved for the indications of complicated intra-abdominal infection (with metronidazole) and complicated urinary tract infection, and have ongoing phase 3 trials for the treatment of ventilator-associated and nosocomial pneumonia. This manuscript will review current data available regarding the spectrum of activity and clinical trials that led to the US Food and Drug Administration (FDA) approval of these agents. Both agents appear to be well tolerated and show promise in the treatment of MDR Gram-negative infections.

  8. A multianalyte ELISA for immunochemical screening of sulfonamide, fluoroquinolone and ss-lactam antibiotics in milk samples using class-selective bioreceptors.

    PubMed

    Adrian, Javier; Pinacho, Daniel G; Granier, Benoit; Diserens, Jean-Marc; Sánchez-Baeza, Francisco; Marco, M-Pilar

    2008-07-01

    A multianalyte ELISA has been developed for the simultaneous determination of the most frequently used antibiotic families in the veterinary field following the typical planar microarray configuration, where the identity of the target analyte is encoded by its location in the detection platform (Master et al. in Drug Discovery Today 11:1007-1011, 2006). To accomplish this aim, two individual enzyme-linked immunosorbent assays for sulfonamide and fluoroquinolone antibiotics and an enzyme-linked receptor assay for ss-lactam antibiotics have been combined. The strategy uses microplates coated with the corresponding haptenized proteins in specific sections of the microplate. The samples are mixed with a cocktail containing the bioreagents, and distributed in the wells of the microplate. Identification of the antibiotic present in a particular sample is consequently accomplished by detecting a positive response on the corresponding microplate section. Since the bioreceptors used show a wide recognition of the congeners of each antibiotic family, the multianalyte method is able to detect more than 25 different antibiotics from the three most important antibiotic families. The detectability reached in full-fat milk samples is below the European maximum residue limits. The accuracy and reliability of this multiplexed bioanalytical method have been demonstrated by analyzing blind spiked samples. PMID:18483810

  9. Reactive extrusion of epsilon-caprolactone polymers and application of poly(lauryl lactam-b-caprolactone) as a compatibilizing agent in blends containing poly(vinyl chloride)

    NASA Astrophysics Data System (ADS)

    Kim, Byong Jun

    In this dissertation, we investigate the continuous polymerization of epsilon-caprolactone (CL) and copolymerization of CL with epsilon-caprolactam (CA), o-lauryl lactam (LA), and styrene (ST) in a modular intermeshing co-rotating twin screw extruder. We consider the variables of temperature profile, screw speed, monomer feed rate, the ratio of monomer to initiator, and feeding order of co-monomers on reactive extrusion of CL polymers. Associated with the reactive extrusion of CL, we also perform the engineering analysis of molecular weight increase and shear-induced molecular weight reduction after polymerization of CL during the reactive extrusion process. Specially designed block copolymers have played a role as compatibilizing agents in the system of immiscible polymer blends. We apply the LA-CL block copolymer (P(LA-b-CL)) produced by reactive extrusion as a compatibilizing agent in immiscible polymer blend systems: (i) poly(vinyl chloride) (PVC)/polyamide 12(PA12), (ii) PVC/polypropylene (PP), and (iii) PVC/maleic anhydride (MA)-modified ethylene-propylene-non-conjugated diene elastomer (MA-EPDM). We investigate the mechanical and thermal properties of (i) PVC/PA12 blend compatibilized with P(LA-b-CL), (ii) PVC/PP blend compatibilized with P(LA-b-CL)/PA12/MA-PP, and (iii) PVC/MA-EPDM blend compatibilized with P(LA-b-CL)/PA12.

  10. Ceftazidime/avibactam: a novel cephalosporin/nonbeta-lactam beta-lactamase inhibitor for the treatment of complicated urinary tract infections and complicated intra-abdominal infections

    PubMed Central

    Hidalgo, Jose A; Vinluan, Celeste M; Antony, Nishaal

    2016-01-01

    There has been greater interest in developing additional antimicrobial agents due to the increasing health care costs and resistance resulting from bacterial pathogens to currently available treatment options. Gram-negative organisms including Enterobacteriaceae and Pseudomonas aeruginosa are some of the most concerning threats due to their resistance mechanisms: extended-spectrum beta-lactamase production and Klebsiella pneumoniae carbapenemase enzymes. Ceftazidime is a third-generation broad-spectrum cephalosporin with activity against P. aeruginosa and avibactam is a novel nonbeta-lactam beta-lactamase inhibitor. Avycaz®, the trade name for this new combination antibiotic, restores the activity of ceftazidime against some of the previously resistant pathogens. Avycaz was approved in 2015 for the treatment of complicated urinary tract infections, including pyelonephritis, and complicated intra-abdominal infections with the addition of metronidazole in patients with little to no other treatment options. This review article assesses the clinical trials and data that led to the approval of this antibiotic, in addition to its spectrum of activity and limitations. PMID:27528799

  11. In vitro activity of beta-lactams, macrolides, telithromycin, and fluoroquinolones against clinical isolates of Streptococcus pneumoniae: correlation between drug resistance and genetic characteristics.

    PubMed

    Yamaguchi, Toshiyuki; Hashikita, Giichi; Takahashi, Shun; Itabashi, Akira; Yamazaki, Tsutomu; Maesaki, Shigefumi

    2005-10-01

    The in vitro activity of antimicrobial agents against Streptococcus pneumoniae was determined using 16 strains of penicillin-susceptible S. pneumoniae (PSSP) and 26 strains of penicillin intermediately resistant S. pneumoniae (PISP) + penicillin-resistant S. pneumoniae (PRSP) in Japan. The minimum inhibitory concentrations (MICs) of potent antibiotics, including eight beta-lactams (benzylpenicillin, ampicillin, cefotiam, cefepime, cefditoren, faropenem, panipenem, and biapenem), three macrolides (erythromycin, clarithromycin, and azithromycin), telithromycin, and three fluoroquinolones (ciprofloxacin, levofloxacin, and gatifloxacin), were determined. Twenty-three strains exhibited genetic variations at pbp1a + pbp2x + pbp2b, which are genetic-PRSP (g-PRSP). g-PISP strains accounted for 62.5% (10/16) of the PSSP strains. The existence of an abnormal pbp gene conferred not only penicillin resistance but resistance to cephems; however, panipenem and biapenem had potent in vitro efficacy against alterations. Regarding the macrolide resistance mechanisms (mefA or ermB): 16 isolates had only mefA, 18 isolates had ermB, and 2 isolates had both mefA and ermB. There was no correlation between the existence of an abnormal pbp gene and the existence of the mefA gene or the ermB gene. PMID:16258826

  12. Exploring the Mechanism of β-Lactam Ring Protonation in the Class A β-lactamase Acylation Mechanism Using Neutron and X-ray Crystallography

    DOE PAGESBeta

    Vandavasi, Venu Gopal; Weiss, Kevin L.; Cooper, Jonathan B.; Erskine, Peter T.; Tomanicek, Stephen J.; Ostermann, Andreas; Schrader, Tobias E.; Ginell, Stephan L.; Coates, Leighton

    2015-12-02

    The catalytic mechanism of class A beta-lactamases is often debated due in part to the large number of amino acids that interact with bound beta-lactam substrates. The role and function of the conserved residue Lys 73 in the catalytic mechanism of class A type beta-lactamase enzymes is still not well understood after decades of scientific research. To better elucidate the functions of this vital residue, we used both neutron and high-resolution X-ray diffraction to examine both the structures of the ligand free protein and the acyl-enzyme complex of perdeuterated E166A Toho-1 beta-lactamase with the antibiotic cefotaxime. The E166A mutant lacksmore » a critical glutamate residue that has a key role in the deacylation step of the catalytic mechanism, allowing the acyl-enzyme adduct to be captured for study. In our ligand free structures, Lys 73 is present in a single conformation, however in all of our acyl-enzyme structures, Lys 73 is present in two different conformations, in which one conformer is closer to Ser 70 while the other conformer is positioned closer to Ser 130, which supports the existence of a possible pathway by which proton transfer from Lys 73 to Ser 130 can occur. This and further clarifications of the role of Lys 73 in the acylation mechanism may facilitate the design of inhibitors that capitalize on the enzymes native machinery.« less

  13. Exploring the Mechanism of β-Lactam Ring Protonation in the Class A β-lactamase Acylation Mechanism Using Neutron and X-ray Crystallography

    SciTech Connect

    Vandavasi, Venu Gopal; Weiss, Kevin L.; Cooper, Jonathan B.; Erskine, Peter T.; Tomanicek, Stephen J.; Ostermann, Andreas; Schrader, Tobias E.; Ginell, Stephan L.; Coates, Leighton

    2015-12-02

    The catalytic mechanism of class A beta-lactamases is often debated due in part to the large number of amino acids that interact with bound beta-lactam substrates. The role and function of the conserved residue Lys 73 in the catalytic mechanism of class A type beta-lactamase enzymes is still not well understood after decades of scientific research. To better elucidate the functions of this vital residue, we used both neutron and high-resolution X-ray diffraction to examine both the structures of the ligand free protein and the acyl-enzyme complex of perdeuterated E166A Toho-1 beta-lactamase with the antibiotic cefotaxime. The E166A mutant lacks a critical glutamate residue that has a key role in the deacylation step of the catalytic mechanism, allowing the acyl-enzyme adduct to be captured for study. In our ligand free structures, Lys 73 is present in a single conformation, however in all of our acyl-enzyme structures, Lys 73 is present in two different conformations, in which one conformer is closer to Ser 70 while the other conformer is positioned closer to Ser 130, which supports the existence of a possible pathway by which proton transfer from Lys 73 to Ser 130 can occur. This and further clarifications of the role of Lys 73 in the acylation mechanism may facilitate the design of inhibitors that capitalize on the enzymes native machinery.

  14. Benzo-fused lactams from a diversity-oriented synthesis (DOS) library as inhibitors of scavenger receptor BI (SR-BI)-mediated lipid uptake.

    PubMed

    Dockendorff, Chris; Faloon, Patrick W; Pu, Jun; Yu, Miao; Johnston, Stephen; Bennion, Melissa; Penman, Marsha; Nieland, Thomas J F; Dandapani, Sivaraman; Perez, José R; Munoz, Benito; Palmer, Michelle A; Schreiber, Stuart L; Krieger, Monty

    2015-01-01

    We report a new series of 8-membered benzo-fused lactams that inhibit cellular lipid uptake from HDL particles mediated by Scavenger Receptor, Class B, Type I (SR-BI). The series was identified via a high-throughput screen of the National Institutes of Health Molecular Libraries Small Molecule Repository (NIH MLSMR), measuring the transfer of the fluorescent lipid DiI from HDL particles to CHO cells overexpressing SR-BI. The series is part of a previously reported diversity-oriented synthesis (DOS) library prepared via a build-couple-pair approach. Detailed structure-activity relationship (SAR) studies were performed with a selection of the original library, as well as additional analogs prepared via solution phase synthesis. These studies demonstrate that the orientation of the substituents on the aliphatic ring have a critical effect on activity. Additionally, a lipophilic group is required at the western end of the molecule, and a northern hydroxyl group and a southern sulfonamide substituent also proved to be optimal. Compound 2p was found to possess a superior combination of potency (av IC50=0.10μM) and solubility (79μM in PBS), and it was designated as probe ML312. PMID:25900219

  15. Novel type of adenylyl cyclase participates in tabtoxinine-β-lactam-induced cell death and occurrence of wildfire disease in Nicotiana benthamiana.

    PubMed

    Ito, Makoto; Takahashi, Hirotaka; Sawasaki, Tatsuya; Ohnishi, Kouhei; Hikichi, Yasufumi; Kiba, Akinori

    2014-01-01

    Tabtoxinine-β-lactam (TβL), a non-specific bacterial toxin, is produced by Pseudomonas syringae pv. tabaci, the causal agent of tobacco wildfire disease. TβL causes the plant cell death by the inhibiting glutamine synthetase, which leads to an abnormal accumulation of ammonium ions. To better understand the molecular mechanisms involved in TβL-induced cell death and necrotic wildfire lesions, we focused on adenylyl cyclase in Nicotiana benthamiana. We isolated the gene designated as NbAC (Nicotiana benthamiana adenylyl cyclase). Recombinant NbAC protein showed adenylyl cyclase activity in vitro. TβL-induced necrotic lesions were significantly suppressed in NbAC-silenced leaves compared with control plant leaves. However, the amount of ammonium ions was scarcely affected by NbAC-silencing. Furthermore, the silencing of NbAC also suppressed l-methionine sulfoximine-induced cell death without any changes in the amount of ammonium accumulated. When inoculated directly with P. syringae pv tabaci, NbAC-silenced plants showed reduced symptoms. These results suggest that NbAC might play an essential role in intracellular signal transduction during TβL-induced cell death and necrotic wildfire disease development.

  16. A possible role for acetylated intermediates in diaminopimelate and tabtoxinine-beta-lactam biosynthesis in Pseudomonas syringae pv. tabaci BR2.024.

    PubMed Central

    Liu, L; Shaw, P D

    1997-01-01

    The deduced product of an open reading frame (ORF3) located in the tabtoxinine-beta-lactam (T beta L) biosynthetic region of Pseudomonas syringae pv. tabaci BR2.024 (BR2.024) has significant sequence homology to the dapD products of other bacteria. dapD encodes L-2,3,4,5-tetrahydrodipicolinate succinyl coenzyme A succinyltransferase (THDPA-ST), an enzyme in the diaminopimelate (DAP) and lysine biosynthetic pathway. Complementation studies, in vitro transcription-translation experiments, and enzymatic assays indicated that ORF3 encodes a product with THDPA-ST activity in Escherichia coli dapD mutant beta 274. However, a BR2.024 mutant with an insert in ORF3 was prototrophic, and only basal THDPA-ST activity was detected in extracts of both parent and mutant. This finding suggested that ORF3 was not required for DAP biosynthesis and that it did not encode a product with THDPA-ST activity. The results of enzymatic studies, indicating that BR2.024 uses acetylated intermediates for DAP biosynthesis, are consistent with the hypothesis that BR2.024 does not need THDPA-ST for DAP biosynthesis. The ORF3 mutant produced reduced levels of tabtoxin, indicating that ORF3 may have a role in T beta L biosynthesis. We have named the gene tabB and have proposed a possible function for the gene product. PMID:9294453

  17. Resolution and Determination of the Absolute Configuration of a Twisted Bis-Lactam Analogue of Tröger's Base: A Comparative Spectroscopic and Computational Study.

    PubMed

    Rúnarsson, Ögmundur Vidar; Benkhäuser, Christian; Christensen, Niels Johan; Ruiz, Josep Artacho; Ascic, Erhad; Harmata, Michael; Snieckus, Victor; Rissanen, Kari; Fristrup, Peter; Lützen, Arne; Wärnmark, Kenneth

    2015-08-21

    The first reported twisted bis-lactam, a racemic Tröger's base (TB) analogue (2), was resolved into its enantiomers on a chiral stationary phase HPLC column. The absolute configuration of (+)-2 was determined to be (R,R)-2 by comparing experimental and calculated vibrational circular dichroism (VCD) and electronic circular dichroism (ECD) spectra. The absolute configuration of (-)-2 was determined by comparing experimental and calculated electronic circular dichroism (ECD) spectra. The corresponding theoretical spectra were calculated using the lowest energy conformation of (R,R)-2 and (S,S)-2 at the B3LYP/6-31G(d,p) level of theory. The absolute configuration of (+)-2 was also determined to (R,R)-2 by anomalous X-ray diffraction (AXRD) in a chiral space group P212121 using Cu-irradiation resulting in a very low Flack parameter of -0.06(3), despite the heaviest element being an oxygen atom, thus unambiguously confirming the results from the spectroscopic studies. We conclude that, for the Tröger's base (TB) analogue (2), we may rank the reliability of the individual methods for AC determination as AXRD ≫ VCD > ECD, while the synergy of all three methods provides very strong confidence in the assigned ACs of (+)-(R,R)-2 and (-)-(S,S)-2. PMID:26244379

  18. HipA-triggered growth arrest and β-lactam tolerance in Escherichia coli are mediated by RelA-dependent ppGpp synthesis.

    PubMed

    Bokinsky, Gregory; Baidoo, Edward E K; Akella, Swetha; Burd, Helcio; Weaver, Daniel; Alonso-Gutierrez, Jorge; García-Martín, Héctor; Lee, Taek Soon; Keasling, Jay D

    2013-07-01

    Persistence is a phenomenon whereby a subpopulation of bacterial cells enters a transient growth-arrested state that confers antibiotic tolerance. While entrance into persistence has been linked to the activities of toxin proteins, the molecular mechanisms by which toxins induce growth arrest and the persistent state remain unclear. Here, we show that overexpression of the protein kinase HipA in Escherichia coli triggers growth arrest by activating synthesis of the alarmone guanosine tetraphosphate (ppGpp) by the enzyme RelA, a signal typically associated with amino acid starvation. We further demonstrate that chemically suppressing ppGpp synthesis with chloramphenicol relieves inhibition of DNA replication initiation and RNA synthesis in HipA-arrested cells and restores vulnerability to β-lactam antibiotics. HipA-arrested cells maintain glucose uptake and oxygen consumption and accumulate amino acids as a consequence of translational inhibition. We harness the active metabolism of HipA-arrested cells to provide a bacteriophage-resistant platform for the production of biotechnologically relevant compounds, which may represent an innovative solution to the costly problem of phage contamination in industrial fermentations.

  19. Antipneumococcal activity of BAY 12-8039, a new quinolone, compared with activities of three other quinolones and four oral beta-lactams.

    PubMed Central

    Visalli, M A; Jacobs, M R; Appelbaum, P C

    1997-01-01

    Activities of BAY 12-8039 against 205 pneumococci were tested by agar dilution. MICs (in micrograms per milliliter) at which 50 and 90% of the isolates are inhibited (MIC50s and MIC90s, respectively) were 0.125 and 0.25 (BAY 12-8039), 2.0 and 4.0 (ciprofloxacin and ofloxacin), and 0.25 and 0.5 (sparfloxacin). Beta-lactam MIC50s and MIC90s for penicillin-susceptible, -intermediate, and -resistant strains, in that order, were 0.016 and 0.03, 0.25 and 2.0, and 2.0 and 4.0 (amoxicillin); 0.03 and 0.06, 0.25 and 4.0, and 4.0 and 8.0 (ampicillin); 0.03 and 0.06, 0.5 and 4.0, and 4.0 and 8.0 (cefuroxime); and 0.03 and 0.125, 0.25 and 2.0, and 4.0 and 8.0 (cefpodoxime). At two times their MICs after 24 h, BAY 12-8039, ciprofloxacin, ampicillin, and cefuroxime were uniformly bactericidal (99.9% killing) against 12 strains; other compounds were bactericidal at four times their MICs. PMID:9420061

  20. New insights into virulence mechanisms of rice pathogen Acidovorax avenae subsp. avenae strain RS-1 following exposure to ß-lactam antibiotics

    PubMed Central

    Li, Bin; Ge, Mengyu; Zhang, Yang; Wang, Li; Ibrahim, Muhammad; Wang, Yanli; Sun, Guochang; Chen, Gongyou

    2016-01-01

    Recent research has shown that pathogen virulence can be altered by exposure to antibiotics, even when the growth rate is unaffected. Investigating this phenomenon provides new insights into understanding the virulence mechanisms of bacterial pathogens. This study investigates the phenotypic and transcriptomic responses of the rice pathogenic bacterium Acidovorax avenae subsp. avenae (Aaa) strain RS-1 to ß-lactam antibiotics especially Ampicillin (Amp). Our results indicate that exposure to Amp does not influence bacterial growth and biofilm formation, but alters the virulence, colonization capacity, composition of extracellular polymeric substances and secretion of Type VI secretion system (T6SS) effector Hcp. This attenuation in virulence is linked to unique or differential expression of known virulence-associated genes based on genome-wide transcriptomic analysis. The reliability of expression data generated by RNA-Seq was verified with quantitative real-time PCR of 21 selected T6SS genes, where significant down-regulation in expression of hcp gene, corresponding to the reduction in secretion of Hcp, was observed under exposure to Amp. Hcp is highlighted as a potential target for Amp, with similar changes observed in virulence-associated phenotypes between exposure to Amp and mutation of hcp gene. In addition, Hcp secretion is reduced in knockout mutants of 4 differentially expressed T6SS genes. PMID:26915352

  1. HipA-Triggered Growth Arrest and β-Lactam Tolerance in Escherichia coli Are Mediated by RelA-Dependent ppGpp Synthesis

    PubMed Central

    Baidoo, Edward E. K.; Akella, Swetha; Burd, Helcio; Weaver, Daniel; Alonso-Gutierrez, Jorge; García-Martín, Héctor; Lee, Taek Soon

    2013-01-01

    Persistence is a phenomenon whereby a subpopulation of bacterial cells enters a transient growth-arrested state that confers antibiotic tolerance. While entrance into persistence has been linked to the activities of toxin proteins, the molecular mechanisms by which toxins induce growth arrest and the persistent state remain unclear. Here, we show that overexpression of the protein kinase HipA in Escherichia coli triggers growth arrest by activating synthesis of the alarmone guanosine tetraphosphate (ppGpp) by the enzyme RelA, a signal typically associated with amino acid starvation. We further demonstrate that chemically suppressing ppGpp synthesis with chloramphenicol relieves inhibition of DNA replication initiation and RNA synthesis in HipA-arrested cells and restores vulnerability to β-lactam antibiotics. HipA-arrested cells maintain glucose uptake and oxygen consumption and accumulate amino acids as a consequence of translational inhibition. We harness the active metabolism of HipA-arrested cells to provide a bacteriophage-resistant platform for the production of biotechnologically relevant compounds, which may represent an innovative solution to the costly problem of phage contamination in industrial fermentations. PMID:23667235

  2. Role of [FeOx(OH)y] surface sites on the adsorption of β-lactamic antibiotics on Al2O3 supported Fe oxide.

    PubMed

    Pinto, Paula S; Medeiros, Tayline P V; Ardisson, José D; Lago, Rochel M

    2016-11-01

    In this work, [FeOx(OH)y]/Al2O3 composites with different Fe oxyhydroxy contents, i.e. 10, 20 and 50wt% treated at 150, 200, 300 and 450°C were investigated as adsorbents of β-lactamic antibiotics, i.e. cephalexin, ceftriaxone and especially amoxicillin, from aqueous solutions. The obtained results showed that the nature of the surface Fe(3+) species play a fundamental role on the adsorption process. The most efficient adsorption was obtained for the sample 150Fe50A (50% [FeOx(OH)y] supported in Al2O3 treated at 150°C) whereas the thermal treatment at higher temperatures caused a strong decrease on the adsorption capacity. Mössbauer, XRD, FTIR, Raman, TG-MS, SEM, CHN and BET of the composite 150Fe50A suggested an approximate composition of FeO0.65(OH)1.7 whereas at 450°C strong dehydroxylation process takes place to form FeO1.4(OH)0.21. These results combined with competitive adsorption using amoxicillin mixed with phosphate or H2O2 suggest that the antibiotic molecules adsorb by complexation on surface sites likely based on FeOx(OH)y by the replacement of the labile OH ligands.

  3. Ceftazidime/avibactam: a novel cephalosporin/nonbeta-lactam beta-lactamase inhibitor for the treatment of complicated urinary tract infections and complicated intra-abdominal infections.

    PubMed

    Hidalgo, Jose A; Vinluan, Celeste M; Antony, Nishaal

    2016-01-01

    There has been greater interest in developing additional antimicrobial agents due to the increasing health care costs and resistance resulting from bacterial pathogens to currently available treatment options. Gram-negative organisms including Enterobacteriaceae and Pseudomonas aeruginosa are some of the most concerning threats due to their resistance mechanisms: extended-spectrum beta-lactamase production and Klebsiella pneumoniae carbapenemase enzymes. Ceftazidime is a third-generation broad-spectrum cephalosporin with activity against P. aeruginosa and avibactam is a novel nonbeta-lactam beta-lactamase inhibitor. Avycaz(®), the trade name for this new combination antibiotic, restores the activity of ceftazidime against some of the previously resistant pathogens. Avycaz was approved in 2015 for the treatment of complicated urinary tract infections, including pyelonephritis, and complicated intra-abdominal infections with the addition of metronidazole in patients with little to no other treatment options. This review article assesses the clinical trials and data that led to the approval of this antibiotic, in addition to its spectrum of activity and limitations. PMID:27528799

  4. Role of [FeOx(OH)y] surface sites on the adsorption of β-lactamic antibiotics on Al2O3 supported Fe oxide.

    PubMed

    Pinto, Paula S; Medeiros, Tayline P V; Ardisson, José D; Lago, Rochel M

    2016-11-01

    In this work, [FeOx(OH)y]/Al2O3 composites with different Fe oxyhydroxy contents, i.e. 10, 20 and 50wt% treated at 150, 200, 300 and 450°C were investigated as adsorbents of β-lactamic antibiotics, i.e. cephalexin, ceftriaxone and especially amoxicillin, from aqueous solutions. The obtained results showed that the nature of the surface Fe(3+) species play a fundamental role on the adsorption process. The most efficient adsorption was obtained for the sample 150Fe50A (50% [FeOx(OH)y] supported in Al2O3 treated at 150°C) whereas the thermal treatment at higher temperatures caused a strong decrease on the adsorption capacity. Mössbauer, XRD, FTIR, Raman, TG-MS, SEM, CHN and BET of the composite 150Fe50A suggested an approximate composition of FeO0.65(OH)1.7 whereas at 450°C strong dehydroxylation process takes place to form FeO1.4(OH)0.21. These results combined with competitive adsorption using amoxicillin mixed with phosphate or H2O2 suggest that the antibiotic molecules adsorb by complexation on surface sites likely based on FeOx(OH)y by the replacement of the labile OH ligands. PMID:27318729

  5. Analysis of Six β-Lactam Residues in Milk and Egg by Micellar Electrokinetic Chromatography with Large-Volume Sample Stacking and Polarity Switching.

    PubMed

    Shao, Yu-Xiu; Chen, Guan-Hua; Fang, Rou; Zhang, Li; Yi, Ling-Xiao; Meng, Hong-Lian

    2016-05-01

    A new micellar electrokinetic chromatography method with large-volume sample stacking and polarity switching was developed to analyze amoxicllin, cephalexin, oxacillin, penicillin G, cefazolin, and cefoperazone in milk and egg. The important parameters influencing separation and enrichment factors were optimized. The optimized running buffer consisted of 10 mM phosphate and 22 mM SDS at pH 6.7. The sample size was 1.47 kPa × 690 s, the reverse voltage was 20 kV, and the electric current recovery was 95%. Under these optimum conditions, the enrichment factors of six β-lactams were 193-601. Their LODs were <0.26 ng/g, and LOQs were all 2 ng/g, which was only 1/50-1/2 of the maximum residual limits demanded by U.S. and Japanese regulations. The intraday and interday RSDs of method were lower than 3.70 and 3.91%, respectively. The method can be applied to determine these six antibiotic residues in egg and milk. PMID:27088652

  6. Development and validation of a sensitive GC-MS method for the determination of trace levels of an alkylating reagent in a beta-lactam active pharmaceutical ingredient.

    PubMed

    Li, Hanlin; Sluggett, Gregory W

    2005-09-15

    A direct injection gas chromatographic method utilizing selected-ion monitoring (SIM) mode mass selective detection was developed and validated for the trace analysis of an impurity, carbonic acid chloromethyl tetrahydro-pyran-4-yl ester (CCMTHP), present in a beta-lactam active pharmaceutical ingredient (API). A variety of analytical techniques including LC-MS, GC-FID, GC-ECD and GC-MS were evaluated during the method development. GC-MS with SIM at m/z=49 demonstrated the best detection sensitivity. A 10 ppm (5 pg on column) limit of quantitation (LOQ) was attained and the linearity of the method was demonstrated in the range of 10-1000 ppm. Accurate and precise quantitation of the impurity in drug substance was achieved with external standardization. A 10:1 split injection was applied to limit the amount of non-volatile API loading onto the column. The effects of injection and detection parameters such as split ratio, liner type, injection temperature and number of mass ions monitored were studied. Full validation proved the accuracy, precision and specificity of the method, which was successfully employed to analyze many pilot lots of the API. PMID:15951149

  7. Abundances of Tetracycline, Sulphonamide and Beta-Lactam Antibiotic Resistance Genes in Conventional Wastewater Treatment Plants (WWTPs) with Different Waste Load

    PubMed Central

    Voolaid, Veiko; Ritz, Christian; Tenson, Tanel; Virta, Marko; Kisand, Veljo

    2014-01-01

    Antibiotics and antibiotic resistant bacteria enter wastewater treatment plants (WWTPs), an environment where resistance genes can potentially spread and exchange between microbes. Several antibiotic resistance genes (ARGs) were quantified using qPCR in three WWTPs of decreasing capacity located in Helsinki, Tallinn, and Tartu, respectively: sulphonamide resistance genes (sul1 and sul2), tetracycline resistance genes (tetM and tetC), and resistance genes for extended spectrum beta-lactams (blaoxa-58, blashv-34, and blactx-m-32). To avoid inconsistencies among qPCR assays we normalised the ARG abundances with 16S rRNA gene abundances while assessing if the respective genes increased or decreased during treatment. ARGs were detected in most samples; sul1, sul2, and tetM were detected in all samples. Statistically significant differences (adjusted p<0.01) between the inflow and effluent were detected in only four cases. Effluent values for blaoxa-58 and tetC decreased in the two larger plants while tetM decreased in the medium-sized plant. Only blashv-34 increased in the effluent from the medium-sized plant. In all other cases the purification process caused no significant change in the relative abundance of resistance genes, while the raw abundances fell by several orders of magnitude. Standard water quality variables (biological oxygen demand, total phosphorus and nitrogen, etc.) were weakly related or unrelated to the relative abundance of resistance genes. Based on our results we conclude that there is neither considerable enrichment nor purification of antibiotic resistance genes in studied conventional WWTPs. PMID:25084517

  8. Novel blaROB-1-Bearing Plasmid Conferring Resistance to β-Lactams in Haemophilus parasuis Isolates from Healthy Weaning Pigs

    PubMed Central

    Moleres, Javier; Santos-López, Alfonso; Lázaro, Isidro; Labairu, Javier; Prat, Cristina; Ardanuy, Carmen; González-Zorn, Bruno

    2015-01-01

    Haemophilus parasuis, the causative agent of Glässer's disease, is one of the early colonizers of the nasal mucosa of piglets. It is prevalent in swine herds, and lesions associated with disease are fibrinous polyserositis and bronchopneumonia. Antibiotics are commonly used in disease control, and resistance to several antibiotics has been described in H. parasuis. Prediction of H. parasuis virulence is currently limited by our scarce understanding of its pathogenicity. Some genes have been associated with H. parasuis virulence, such as lsgB and group 1 vtaA, while biofilm growth has been associated with nonvirulent strains. In this study, 86 H. parasuis nasal isolates from farms that had not had a case of disease for more than 10 years were obtained by sampling piglets at weaning. Isolates were studied by enterobacterial repetitive intergenic consensus PCR and determination of the presence of lsgB and group 1 vtaA, biofilm formation, inflammatory cell response, and resistance to antibiotics. As part of the diversity encountered, a novel 2,661-bp plasmid, named pJMA-1, bearing the blaROB-1 β-lactamase was detected in eight colonizing strains. pJMA-1 was shown to share a backbone with other small plasmids described in the Pasteurellaceae, to be 100% stable, and to have a lower biological cost than the previously described plasmid pB1000. pJMA-1 was also found in nine H. parasuis nasal strains from a separate collection, but it was not detected in isolates from the lesions of animals with Glässer's disease or in nontypeable Haemophilus influenzae isolates. Altogether, we show that commensal H. parasuis isolates represent a reservoir of β-lactam resistance genes which can be transferred to pathogens or other bacteria. PMID:25747001

  9. Impaired mitochondrial function in HepG2 cells treated with hydroxy-cobalamin[c-lactam]: A cell model for idiosyncratic toxicity.

    PubMed

    Haegler, Patrizia; Grünig, David; Berger, Benjamin; Krähenbühl, Stephan; Bouitbir, Jamal

    2015-10-01

    The vitamin B12 analog hydroxy-cobalamin[c-lactam] (HCCL) impairs mitochondrial protein synthesis and the function of the electron transport chain. Our goal was to establish an in vitro model for mitochondrial dysfunction in human hepatoma cells (HepG2), which can be used to investigate hepatotoxicity of idiosyncratic mitochondrial toxicants. For that, HepG2 cells were treated with HCCL, which inhibits the function of methylmalonyl-CoA mutase and impairs mitochondrial protein synthesis. Secondary, cells were incubated with propionate that served as source of propionyl-CoA, a percursor of methylmalonyl-CoA. Dose-finding experiments were conducted to evaluate the optimal dose and treatment time of HCCL and propionate for experiments on mitochondrial function. 50 μM HCCL was cytotoxic after exposure of HepG2 cells for 2d and 10 and 50 μM HCCL enhanced the cytotoxicity of 100 or 1000 μM propionate. Co-treatment with HCCL (10 μM) and propionate (1000 μM) dissipated the mitochondrial membrane potential and impaired the activity of enzyme complex IV of the electron transport chain. Treatment with HCCL decreased the mRNA content of mitochondrially encoded proteins, whereas the mtDNA content remained unchanged. We observed mitochondrial ROS accumulation and decreased mitochondrial SOD2 expression. Moreover, electron microscopy showed mitochondrial swelling. Finally, HepG2 cells pretreated with a non-cytotoxic combination of HCCL (10 μM) and propionate (100 μM) were more sensitive to the mitochondrial toxicants dronedarone, benzbromarone, and ketoconazole than untreated cells. In conclusion, we established and characterized a cell model, which could be used for testing drugs with idiosyncratic mitochondrial toxicity.

  10. New hydrolysis products of the beta-lactam antibiotic amoxicillin, their pH-dependent formation and search in municipal wastewater.

    PubMed

    Hirte, Kristin; Seiwert, Bettina; Schüürmann, Gerrit; Reemtsma, Thorsten

    2016-01-01

    Amoxicillin (AMX) is a widespread β-lactam-antibiotic and, together with some of its transformation products (TPs) originating from hydrolysis, a known environmental contaminant. To shed light on the abiotic degradation of AMX and the stability of its known TPs, laboratory hydrolysis experiments of AMX were carried out at pH 3, 7 and 11. Not only the rate of hydrolysis but also the pattern of TPs was strongly pH-dependent. The time courses of the obtained transformation products were analyzed by UPLC-HR-QToF-MS. AMX penicilloic acid (TP 1), AMX 2',5'-diketopiperazine (TP 2), AMX penilloic acid (TP 3) and 3-(4-hydroxyphenyl)pyrazinol (TP 4) were found at neutral pH. Surprisingly, the first three were not stable but transformed into 23 yet unknown TPs within three to four weeks. Seven TPs were tentatively identified, based on their product ion spectra and, where possible, confirmed with reference standards, e.g. penicillamine disulfide, 2-[amino(carboxy)methyl]-5,5-dimethyl-1,3-thiazolidine-4-carboxylic acid and dehydrocarboxylated amoxicillin penilloic acid. Analysis of samples from municipal wastewater treatment plants confirmed these findings with TP 1 being the dominant TP in the influent and a shift towards TP 2, TP 3 and TP 4 in the effluents. The lab experiments predicted up to 13 consecutive TPs from TP 1, TP 2 and TP 3 under neutral conditions. Their detection from surface waters will be difficult, because their large number and slow formation kinetics will lead to comparatively low environmental concentrations. Nevertheless the abiotic degradation of TP 1, TP 2 and TP 3 to further TPs needs to be considered in future studies of the environmental fate of amoxicillin.

  11. Abundances of tetracycline, sulphonamide and beta-lactam antibiotic resistance genes in conventional wastewater treatment plants (WWTPs) with different waste load.

    PubMed

    Laht, Mailis; Karkman, Antti; Voolaid, Veiko; Ritz, Christian; Tenson, Tanel; Virta, Marko; Kisand, Veljo

    2014-01-01

    Antibiotics and antibiotic resistant bacteria enter wastewater treatment plants (WWTPs), an environment where resistance genes can potentially spread and exchange between microbes. Several antibiotic resistance genes (ARGs) were quantified using qPCR in three WWTPs of decreasing capacity located in Helsinki, Tallinn, and Tartu, respectively: sulphonamide resistance genes (sul1 and sul2), tetracycline resistance genes (tetM and tetC), and resistance genes for extended spectrum beta-lactams (blaoxa-58, blashv-34, and blactx-m-32). To avoid inconsistencies among qPCR assays we normalised the ARG abundances with 16S rRNA gene abundances while assessing if the respective genes increased or decreased during treatment. ARGs were detected in most samples; sul1, sul2, and tetM were detected in all samples. Statistically significant differences (adjusted p<0.01) between the inflow and effluent were detected in only four cases. Effluent values for blaoxa-58 and tetC decreased in the two larger plants while tetM decreased in the medium-sized plant. Only blashv-34 increased in the effluent from the medium-sized plant. In all other cases the purification process caused no significant change in the relative abundance of resistance genes, while the raw abundances fell by several orders of magnitude. Standard water quality variables (biological oxygen demand, total phosphorus and nitrogen, etc.) were weakly related or unrelated to the relative abundance of resistance genes. Based on our results we conclude that there is neither considerable enrichment nor purification of antibiotic resistance genes in studied conventional WWTPs.

  12. Characterization of a Novel Small Molecule That Potentiates β-Lactam Activity against Gram-Positive and Gram-Negative Pathogens

    PubMed Central

    Nair, Dhanalakshmi R.; Monteiro, João M.; Memmi, Guido; Thanassi, Jane; Pucci, Michael; Schwartzman, Joseph; Pinho, Mariana G.

    2015-01-01

    In a loss-of-viability screen using small molecules against methicillin-resistant Staphylococcus aureus (MRSA) strain USA300 with a sub-MIC of a β-lactam, we found a small molecule, designated DNAC-1, which potentiated the effect of oxacillin (i.e., the MIC of oxacillin decreased from 64 to 0.25 μg/ml). Fluorescence microscopy indicated a disruption in the membrane structures within 15 min of exposure to DNAC-1 at 2× MIC. This permeabilization was accompanied by a rapid loss of membrane potential, as monitored by use of the DiOC2 (3,3′-diethyloxacarbocyanine iodide) dye. Macromolecular analysis showed the inhibition of staphylococcal cell wall synthesis by DNAC-1. Transmission electron microscopy of treated MRSA USA300 cells revealed a slightly thicker cell wall, together with mesosome-like projections into the cytosol. The exposure of USA300 cells to DNAC-1 was associated with the mislocalization of FtsZ accompanied by the localization of penicillin-binding protein 2 (PBP2) and PBP4 away from the septum, as well as mild activation of the vraRS-mediated cell wall stress response. However, DNAC-1 does not have any generalized toxicity toward mammalian host cells. DNAC-1 in combination with ceftriaxone is also effective against an assortment of Gram-negative pathogens. Using a murine subcutaneous coinjection model with 108 CFU of USA300 as a challenge inoculum, DNAC-1 alone or DNAC-1 with a sub-MIC of oxacillin resulted in a 6-log reduction in bacterial load and decreased abscess formation compared to the untreated control. We propose that DNAC-1, by exerting a bimodal effect on the cell membrane and cell wall, is a viable candidate in the development of combination therapy against many common bacterial pathogens. PMID:25583731

  13. Beneficial Antimicrobial Effect of the Addition of an Aminoglycoside to a β-Lactam Antibiotic in an E. coli Porcine Intensive Care Severe Sepsis Model

    PubMed Central

    Skorup, Paul; Maudsdotter, Lisa; Lipcsey, Miklós; Castegren, Markus; Larsson, Anders; Jonsson, Ann-Beth; Sjölin, Jan

    2014-01-01

    This study aimed to determine whether the addition of an aminoglycoside to a ß-lactam antibiotic increases the antimicrobial effect during the early phase of Gram-negative severe sepsis/septic shock. A porcine model was selected that considered each animal’s individual blood bactericidal capacity. Escherichia coli, susceptible to both antibiotics, was given to healthy pigs intravenously during 3 h. At 2 h, the animals were randomized to a 20-min infusion with either cefuroxime alone (n = 9), a combination of cefuroxime+tobramycin (n = 9), or saline (control, n = 9). Blood samples were collected hourly for cultures and quantitative polymerase chain reaction (PCR). Bacterial growth in the organs after 6 h was chosen as the primary endpoint. A blood sample was obtained at baseline before start of bacterial infusion for ex vivo investigation of the blood bactericidal capacity. At 1 h after the administration of the antibiotics, a second blood sample was taken for ex vivo investigation of the antibiotic-induced blood killing activity. All animals developed severe sepsis/septic shock. Blood cultures and PCR rapidly became negative after completed bacterial infusion. Antibiotic-induced blood killing activity was significantly greater in the combination group than in the cefuroxime group (p<0.001). Growth of bacteria in the spleen was reduced in the two antibiotic groups compared with the controls (p<0.01); no difference was noted between the two antibiotic groups. Bacterial growth in the liver was significantly less in the combination group than in the cefuroxime group (p<0.05). High blood bactericidal capacity at baseline was associated with decreased growth in the blood and spleen (p<0.05). The addition of tobramycin to cefuroxime results in increased antibiotic-induced blood killing activity and less bacteria in the liver than cefuroxime alone. Individual blood bactericidal capacity may have a significant effect on antimicrobial outcome. PMID:24587365

  14. Epicatechin gallate, a naturally occurring polyphenol, alters the course of infection with β-lactam-resistant Staphylococcus aureus in the zebrafish embryo

    PubMed Central

    Stevens, Christina S.; Rosado, Helena; Harvey, Robert J.; Taylor, Peter W.

    2015-01-01

    (-)-epicatechin gallate (ECg) substantially modifies the properties of Staphylococcus aureus and reversibly abrogates β-lactam resistance in methicillin/oxacillin resistant (MRSA) isolates. We have determined the capacity of ECg to alter the course of infection in zebrafish embryos challenged with epidemic clinical isolate EMRSA-16. At 30 h post fertilization (hpf), embryos were infected by injection of 1–5 × 103 colony forming units (CFU) of EMRSA-16 into the circulation valley or yolk sac. Infection by yolk sac injection was lethal with a challenge dose above 3 × 103 CFU, with no survivors at 70 hpf. In contrast, survival at 70 hpf after injection into the circulation was 83 and 44% following challenge with 3 × 103 and 1–5 × 103 CFU, respectively. No significant increases in survival were noted when infected embryos were maintained in medium containing 12.5–100 μg/mL ECg with or without 4 or 16 μg/mL oxacillin. However, when EMRSA-16 was grown in medium containing 12.5 μg/mL ECg and the bacteria used to infect embryos by either the circulation valley or yolk sac, there were significant increases in embryo survival in both the presence and absence of oxacillin. ECg-modified and unmodified, GFP-transformed EMRSA-16 bacteria were visualized within phagocytic cells in the circulation and yolk sac; pre-treatment with ECg also significantly increased induction of the respiratory burst and suppressed increases in IL-1β expression typical of infection with untreated EMRSA-16. We conclude that exposure to ECg prior to infection reduces the lethality of EMRSA-16, renders cells more susceptible to elimination by immune processes and compromises their capacity to establish an inflammatory response in comparison to non-exposed bacteria. PMID:26441953

  15. Investigation of the influence of different physico-chemical parameters upon the susceptibility of planktonic and adherent Escherichia coli cells to beta-lactams and quinolones.

    PubMed

    Drăcea, O; Iordache, C; Bucur, M; Bleotu, C; Banu, O; Ungureanu, C; Cristea, D; Lixandru, M S; Larion, Cristina; Necula, G; Lazăr, V; Chifiriuc, M C

    2009-01-01

    The purpose of this study was to evaluate the influence of different physico-chemical parameters on Escherichia coli susceptibility to ceftriaxone (CRO), cefotaxime (CTX), imipenem (IMP), and nalidixic acid (as marker for resistance by impermeability). The influence of chemical composition of culture medium was evaluated by the comparative assessment of inhibition growth diameters on different solid media: Mueller Hinton Medium (MH), Plate Count Agar Medium (PCA), MacConkey Medium (MC) and Eosin Methylen Blue Medium (EMB). In order to evaluate the differences in antibiotic susceptibility between the biofilm embedded and planktonic cells, an original, simple experimental model was used, by including the bacterial cells in an agar layer, mimicking the biofilm matrix. Our results demonstrated that the inhibition diameter zone was much larger on PCA, EMB and MC than on MH, considered as general standard medium for the antibiosusceptibility testings (CLSI). When bacterial cells were included in the agar matrix, the growth inhibition diameters obtained for different beta-lactams proved to be different of planktonic cells, i.e.: for CTX, a narrow inhibition diameter was obtained, demonstrating the low efficiency of this antibiotic in the treatment of biofilm associated infections, whereas the CRO proved the same efficiency against planktonic as well as to agar embedded bacteria. The different susceptibility results obtained for the cells embedded in the agar matrix by an adapted disk diffusion method are pleading for the necessity to assess new adapted standard methods and specific parameters in the purpose to determine the antibiotic resistance of bacterial cells isolated from biofilm associated infections.

  16. Characterization of a novel small molecule that potentiates β-lactam activity against gram-positive and gram-negative pathogens.

    PubMed

    Nair, Dhanalakshmi R; Monteiro, João M; Memmi, Guido; Thanassi, Jane; Pucci, Michael; Schwartzman, Joseph; Pinho, Mariana G; Cheung, Ambrose L

    2015-04-01

    In a loss-of-viability screen using small molecules against methicillin-resistant Staphylococcus aureus (MRSA) strain USA300 with a sub-MIC of a β-lactam, we found a small molecule, designated DNAC-1, which potentiated the effect of oxacillin (i.e., the MIC of oxacillin decreased from 64 to 0.25 μg/ml). Fluorescence microscopy indicated a disruption in the membrane structures within 15 min of exposure to DNAC-1 at 2× MIC. This permeabilization was accompanied by a rapid loss of membrane potential, as monitored by use of the DiOC2 (3,3'-diethyloxacarbocyanine iodide) dye. Macromolecular analysis showed the inhibition of staphylococcal cell wall synthesis by DNAC-1. Transmission electron microscopy of treated MRSA USA300 cells revealed a slightly thicker cell wall, together with mesosome-like projections into the cytosol. The exposure of USA300 cells to DNAC-1 was associated with the mislocalization of FtsZ accompanied by the localization of penicillin-binding protein 2 (PBP2) and PBP4 away from the septum, as well as mild activation of the vraRS-mediated cell wall stress response. However, DNAC-1 does not have any generalized toxicity toward mammalian host cells. DNAC-1 in combination with ceftriaxone is also effective against an assortment of Gram-negative pathogens. Using a murine subcutaneous coinjection model with 10(8) CFU of USA300 as a challenge inoculum, DNAC-1 alone or DNAC-1 with a sub-MIC of oxacillin resulted in a 6-log reduction in bacterial load and decreased abscess formation compared to the untreated control. We propose that DNAC-1, by exerting a bimodal effect on the cell membrane and cell wall, is a viable candidate in the development of combination therapy against many common bacterial pathogens.

  17. Structural Variabilities in β-Lactamase (blaA) of Different Biovars of Yersinia enterocolitica: Implications for β-Lactam Antibiotic and β-Lactamase Inhibitor Susceptibilities

    PubMed Central

    Singhal, Neelja; Srivastava, Abhishikha; Kumar, Manish; Virdi, Jugsharan Singh

    2015-01-01

    Yersiniosis caused by Yersinia enterocolitica has been reported from all continents. The bacterial species is divided into more than fifty serovars and six biovars viz. 1A, 1B, 2, 3, 4 and 5 which differ in geographical distribution, ecological niches and pathogenicity. Most Y.enterocolitica strains harbor chromosomal genes for two β-lactamases, blaA an Ambler class A penicillinase and blaB an Ambler class C inducible cephalosporinase. In the present study, susceptibility to b-lactam antibiotics and β-lactamase inhibitor was studied for Y. enterocolitica strains of biovars 1A, 1B, 2 and 4. We observed that β-lactamases were expressed differentially among strains of different biovars. To understand the molecular mechanisms underlying such differential expression, the sequences of genes and promoters of blaA were compared. Also, the variants of blaA present in different biovars were modeled and docked with amoxicillin and clavulanic acid. The mRNA secondary structures of blaA variants were also predicted in-silico. Our findings indicated that neither variations in the promoter regions, nor the secondary structures of mRNA contributed to higher/lower expression of blaA in different biovars. Analysis of H-bonding residues of blaA variants with amoxicillin and clavulanic acid revealed that if amino acid residues of a β-lactamase interacting with amoxicillin and the clavulanic acid were similar, clavulanic acid was effective in engaging the enzyme, accounting for a significant reduction in MIC of amoxicillin-clavulanate. This finding might aid in designing better β-lactamase inhibitors with improved efficiencies in future. PMID:25919756

  18. Crystal Structure of the Extended-Spectrum β-Lactamase PER-2 and Insights into the Role of Specific Residues in the Interaction with β-Lactams and β-Lactamase Inhibitors

    PubMed Central

    Ruggiero, Melina; Kerff, Frédéric; Herman, Raphaël; Sapunaric, Frédéric; Galleni, Moreno; Gutkind, Gabriel; Charlier, Paulette; Sauvage, Eric

    2014-01-01

    PER-2 belongs to a small (7 members to date) group of extended-spectrum β-lactamases. It has 88% amino acid identity with PER-1 and both display high catalytic efficiencies toward most β-lactams. In this study, we determined the X-ray structure of PER-2 at 2.20 Å and evaluated the possible role of several residues in the structure and activity toward β-lactams and mechanism-based inhibitors. PER-2 is defined by the presence of a singular trans bond between residues 166 to 167, which generates an inverted Ω loop, an expanded fold of this domain that results in a wide active site cavity that allows for efficient hydrolysis of antibiotics like the oxyimino-cephalosporins, and a series of exclusive interactions between residues not frequently involved in the stabilization of the active site in other class A β-lactamases. PER β-lactamases might be included within a cluster of evolutionarily related enzymes harboring the conserved residues Asp136 and Asn179. Other signature residues that define these enzymes seem to be Gln69, Arg220, Thr237, and probably Arg/Lys240A (“A” indicates an insertion according to Ambler's scheme for residue numbering in PER β-lactamases), with structurally important roles in the stabilization of the active site and proper orientation of catalytic water molecules, among others. We propose, supported by simulated models of PER-2 in combination with different β-lactams, the presence of a hydrogen-bond network connecting Ser70-Gln69-water-Thr237-Arg220 that might be important for the proper activity and inhibition of the enzyme. Therefore, we expect that mutations occurring in these positions will have impacts on the overall hydrolytic behavior. PMID:25070104

  19. Interspecies Transfer of the Penicillin-Binding Protein 3-Encoding Gene ftsI between Haemophilus influenzae and Haemophilus haemolyticus Can Confer Reduced Susceptibility to β-Lactam Antimicrobial Agents

    PubMed Central

    Søndergaard, Annette; Witherden, Elizabeth A.

    2015-01-01

    Mutations in ftsI, encoding penicillin-binding protein 3, can cause decreased β-lactam susceptibility in Haemophilus influenzae. Sequencing of ftsI from clinical strains has indicated interspecies recombination of ftsI between H. influenzae and Haemophilus haemolyticus. This study documented apparently unrestricted homologous recombination of ftsI between H. influenzae and H. haemolyticus in vitro. Transfer of ftsI from resistant isolates conferred similar but not identical increases in the MICs of susceptible strains of H. influenzae and H. haemolyticus. PMID:25918135

  20. A Multinational, Preregistered Cohort Study of β-Lactam/β-Lactamase Inhibitor Combinations for Treatment of Bloodstream Infections Due to Extended-Spectrum-β-Lactamase-Producing Enterobacteriaceae.

    PubMed

    Gutiérrez-Gutiérrez, Belén; Pérez-Galera, Salvador; Salamanca, Elena; de Cueto, Marina; Calbo, Esther; Almirante, Benito; Viale, Pierluigi; Oliver, Antonio; Pintado, Vicente; Gasch, Oriol; Martínez-Martínez, Luis; Pitout, Johann; Akova, Murat; Peña, Carmen; Molina, José; Hernández, Alicia; Venditti, Mario; Prim, Nuria; Origüen, Julia; Bou, German; Tacconelli, Evelina; Tumbarello, Mario; Hamprecht, Axel; Giamarellou, Helen; Almela, Manel; Pérez, Federico; Schwaber, Mitchell J; Bermejo, Joaquín; Lowman, Warren; Hsueh, Po-Ren; Mora-Rillo, Marta; Natera, Clara; Souli, Maria; Bonomo, Robert A; Carmeli, Yehuda; Paterson, David L; Pascual, Alvaro; Rodríguez-Baño, Jesús

    2016-07-01

    The spread of extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae (ESBL-E) is leading to increased carbapenem consumption. Alternatives to carbapenems need to be investigated. We investigated whether β-lactam/β-lactamase inhibitor (BLBLI) combinations are as effective as carbapenems in the treatment of bloodstream infections (BSI) due to ESBL-E. A multinational, retrospective cohort study was performed. Patients with monomicrobial BSI due to ESBL-E were studied; specific criteria were applied for inclusion of patients in the empirical-therapy (ET) cohort (ETC; 365 patients), targeted-therapy (TT) cohort (TTC; 601 patients), and global cohort (GC; 627 patients). The main outcome variables were cure/improvement rate at day 14 and all-cause 30-day mortality. Multivariate analysis, propensity scores (PS), and sensitivity analyses were used to control for confounding. The cure/improvement rates with BLBLIs and carbapenems were 80.0% and 78.9% in the ETC and 90.2% and 85.5% in the TTC, respectively. The 30-day mortality rates were 17.6% and 20% in the ETC and 9.8% and 13.9% in the TTC, respectively. The adjusted odds ratio (OR) (95% confidence interval [CI]) values for cure/improvement rate with ET with BLBLIs were 1.37 (0.69 to 2.76); for TT, they were 1.61 (0.58 to 4.86). Regarding 30-day mortality, the adjusted OR (95% CI) values were 0.55 (0.25 to 1.18) for ET and 0.59 (0.19 to 1.71) for TT. The results were consistent in all subgroups studied, in a stratified analysis according to quartiles of PS, in PS-matched cases, and in the GC. BLBLIs, if active in vitro, appear to be as effective as carbapenems for ET and TT of BSI due to ESLB-E regardless of the source and specific species. These data may help to avoid the overuse of carbapenems. (This study has been registered at ClinicalTrials.gov under registration no. NCT01764490.). PMID:27139473

  1. Fosfomycin plus β-Lactams as Synergistic Bactericidal Combinations for Experimental Endocarditis Due to Methicillin-Resistant and Glycopeptide-Intermediate Staphylococcus aureus.

    PubMed

    del Río, A; García-de-la-Mària, C; Entenza, J M; Gasch, O; Armero, Y; Soy, D; Mestres, C A; Pericás, J M; Falces, C; Ninot, S; Almela, M; Cervera, C; Gatell, J M; Moreno, A; Moreillon, P; Marco, F; Miró, J M

    2016-01-01

    The urgent need of effective therapies for methicillin-resistant Staphylococcus aureus (MRSA) infective endocarditis (IE) is a cause of concern. We aimed to ascertain the in vitro and in vivo activity of the older antibiotic fosfomycin combined with different beta-lactams against MRSA and glycopeptide-intermediate-resistant S. aureus (GISA) strains. Time-kill tests with 10 isolates showed that fosfomycin plus imipenem (FOF+IPM) was the most active evaluated combination. In an aortic valve IE model with two strains (MRSA-277H and GISA-ATCC 700788), the following intravenous regimens were compared: fosfomycin (2 g every 8 h [q8h]) plus imipenem (1 g q6h) or ceftriaxone (2 g q12h) (FOF+CRO) and vancomycin at a standard dose (VAN-SD) (1 g q12h) and a high dose (VAN-HD) (1 g q6h). Whereas a significant reduction of MRSA-227H load in the vegetations (veg) was observed with FOF+IPM compared with VAN-SD (0 [interquartile range [IQR], 0 to 1] versus 2 [IQR, 0 to 5.1] log CFU/g veg; P = 0.01), no statistical differences were found with VAN-HD. In addition, FOF+IPM sterilized more vegetations than VAN-SD (11/15 [73%] versus 5/16 [31%]; P = 0.02). The GISA-ATCC 700788 load in the vegetations was significantly lower after FOF+IPM or FOF+CRO treatment than with VAN-SD (2 [IQR, 0 to 2] and 0 [IQR, 0 to 2] versus 6.5 [IQR, 2 to 6.9] log CFU/g veg; P < 0.01). The number of sterilized vegetations after treatment with FOF+CRO was higher than after treatment with VAN-SD or VAN-HD (8/15 [53%] versus 4/20 [20%] or 4/20 [20%]; P = 0.03). To assess the effect of FOF+IPM on penicillin binding protein (PBP) synthesis, molecular studies were performed, with results showing that FOF+IPM treatment significantly decreased PBP1, PBP2 (but not PBP2a), and PBP3 synthesis. These results allow clinicians to consider the use of FOF+IPM or FOF+CRO to treat MRSA or GISA IE. PMID:26525803

  2. β-Amino acids containing peptides and click-cyclized peptide as β-turn mimics: a comparative study with 'conventional' lactam- and disulfide-bridged hexapeptides.

    PubMed

    Larregola, Maud; Lequin, Olivier; Karoyan, Philippe; Guianvarc'h, Dominique; Lavielle, Solange

    2011-09-01

    The increasing interest in click chemistry and its use to stabilize turn structures led us to compare the propensity for β-turn stabilization of different analogs designed as mimics of the β-turn structure found in tendamistat. The β-turn conformation of linear β-amino acid-containing peptides and triazole-cyclized analogs were compared to 'conventional' lactam- and disulfide-bridged hexapeptide analogs. Their 3D structures and their propensity to fold in β-turns in solution, and for those not structured in solution in the presence of α-amylase, were analyzed by NMR spectroscopy and by restrained molecular dynamics with energy minimization. The linear tetrapeptide Ac-Ser-Trp-Arg-Tyr-NH(2) and both the amide bond-cyclized, c[Pro-Ser-Trp-Arg-Tyr-D-Ala] and the disulfide-bridged, Ac-c[Cys-Ser-Trp-Arg-Tyr-Cys]-NH(2) hexapeptides adopt dominantly in solution a β-turn conformation closely related to the one observed in tendamistat. On the contrary, the β-amino acid-containing peptides such as Ac-(R)-β(3) -hSer-(S)-Trp-(S)-β(3) -hArg-(S)-β(3) -hTyr-NH(2) , and the triazole cyclic peptide, c[Lys-Ser-Trp-Arg-Tyr-βtA]-NH(2) , both specifically designed to mimic this β-turn, do not adopt stable structures in solution and do not show any characteristics of β-turn conformation. However, these unstructured peptides specifically interact in the active site of α-amylase, as shown by TrNOESY and saturation transfer difference NMR experiments performed in the presence of the enzyme, and are displaced by acarbose, a specific α-amylase inhibitor. Thus, in contrast to amide-cyclized or disulfide-bridged hexapeptides, β-amino acid-containing peptides and click-cyclized peptides may not be regarded as β-turn stabilizers, but can be considered as potential β-turn inducers.

  3. Fosfomycin plus β-Lactams as Synergistic Bactericidal Combinations for Experimental Endocarditis Due to Methicillin-Resistant and Glycopeptide-Intermediate Staphylococcus aureus

    PubMed Central

    del Río, A.; García-de-la-Mària, C.; Entenza, J. M.; Gasch, O.; Armero, Y.; Soy, D.; Mestres, C. A.; Pericás, J. M.; Falces, C.; Ninot, S.; Almela, M.; Cervera, C.; Gatell, J. M.; Moreno, A.; Moreillon, P.; Marco, F.

    2015-01-01

    The urgent need of effective therapies for methicillin-resistant Staphylococcus aureus (MRSA) infective endocarditis (IE) is a cause of concern. We aimed to ascertain the in vitro and in vivo activity of the older antibiotic fosfomycin combined with different beta-lactams against MRSA and glycopeptide-intermediate-resistant S. aureus (GISA) strains. Time-kill tests with 10 isolates showed that fosfomycin plus imipenem (FOF+IPM) was the most active evaluated combination. In an aortic valve IE model with two strains (MRSA-277H and GISA-ATCC 700788), the following intravenous regimens were compared: fosfomycin (2 g every 8 h [q8h]) plus imipenem (1 g q6h) or ceftriaxone (2 g q12h) (FOF+CRO) and vancomycin at a standard dose (VAN-SD) (1 g q12h) and a high dose (VAN-HD) (1 g q6h). Whereas a significant reduction of MRSA-227H load in the vegetations (veg) was observed with FOF+IPM compared with VAN-SD (0 [interquartile range [IQR], 0 to 1] versus 2 [IQR, 0 to 5.1] log CFU/g veg; P = 0.01), no statistical differences were found with VAN-HD. In addition, FOF+IPM sterilized more vegetations than VAN-SD (11/15 [73%] versus 5/16 [31%]; P = 0.02). The GISA-ATCC 700788 load in the vegetations was significantly lower after FOF+IPM or FOF+CRO treatment than with VAN-SD (2 [IQR, 0 to 2] and 0 [IQR, 0 to 2] versus 6.5 [IQR, 2 to 6.9] log CFU/g veg; P < 0.01). The number of sterilized vegetations after treatment with FOF+CRO was higher than after treatment with VAN-SD or VAN-HD (8/15 [53%] versus 4/20 [20%] or 4/20 [20%]; P = 0.03). To assess the effect of FOF+IPM on penicillin binding protein (PBP) synthesis, molecular studies were performed, with results showing that FOF+IPM treatment significantly decreased PBP1, PBP2 (but not PBP2a), and PBP3 synthesis. These results allow clinicians to consider the use of FOF+IPM or FOF+CRO to treat MRSA or GISA IE. PMID:26525803

  4. A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface.

    PubMed

    Burke, J R; Gregor, K R; Padmanabha, R; Banville, J; Witmer, M R; Davern, L B; Manly, S P; Tramposch, K M

    1998-06-01

    Cytosolic phospholipase A2 (cPLA2) catalyzes the selective release of arachidonic acid from the sn-2 position of phospholipids and is believed to play a key cellular role in the generation of arachidonic acid. When assaying the human recombinant cPLA2 using membranes isolated from [3H]arachidonate-labeled U937 cells as substrate, 3,3-Dimethyl-6-(3-lauroylureido)-7-oxo-4-thia-1-azabicyclo[3,2,0] heptane-2-carboxylic acid (1) was found to inhibit the enzyme in a dose-dependent manner (IC50 = 72 microM). This beta-lactam did not inhibit other phospholipases, including the human nonpancreatic secreted phospholipase A2. The inhibition of cPLA2 was found not to be time-dependent. This, along with the observation that the degradation of the inhibitor was not catalyzed by the enzyme, demonstrates that the inhibition does not result from the formation of an acyl-enzyme intermediate with the active site serine residue. Moreover, the ring-opened form of 1 is also able to inhibit cPLA2 with near-equal potency. To further characterize the mechanism of inhibition, an assay in which the enzyme is bound to vesicles of 1,2-dimyristoyl-sn-glycero-3-phosphomethanol containing 6-10 mole percent of 1-palmitoyl-2-[1-14C]-arachidonoyl-sn-glycero-3-phosphocholine was employed. With this substrate system, the dose-dependent inhibition was defined by kinetic equations describing competitive inhibition at the lipid/water interface. The apparent dissociation constant for the inhibitor bound to the enzyme at the interface (KI*app) was determined to be 0.5 +/- 0.1 mole% versus an apparent dissociation constant for the arachidonate-containing phospholipid of 0.4 +/- 0.1 mole%. Thus, 1 represents a novel structural class of inhibitors of cPLA2 which partitions into the phospholipid bilayer and competes with the phospholipid substrate for the active site.

  5. Discovery of further pyrrolidine trans-lactams as inhibitors of human neutrophil elastase (HNE) with potential as development candidates and the crystal structure of HNE complexed with an inhibitor (GW475151).

    PubMed

    Macdonald, Simon J F; Dowle, Michael D; Harrison, Lee A; Clarke, Geoffrey D E; Inglis, Graham G A; Johnson, Martin R; Shah, Pritom; Smith, Robin A; Amour, Augustin; Fleetwood, Gill; Humphreys, Davina C; Molloy, Christopher R; Dixon, Mary; Godward, Rosalind E; Wonacott, Alan J; Singh, Onkar M P; Hodgson, Simon T; Hardy, George W

    2002-08-29

    Described herein is a modern approach to the rapid preparation and evaluation of compounds as potential back-up drug candidates. GW311616A, 1, a derivative of pyrrolidine trans-lactams, has previously been described as a potent, orally active inhibitor of human neutrophil elastase (HNE) for the treatment of respiratory disease. These properties made it a suitable candidate for development. Described here is the discovery of three further derivatives of pyrrolidine trans-lactams, which fulfill the criteria required for back-up candidates 28, 29, and 32. These include increased activity in inhibiting HNE in human whole blood (HWB) and comparable pharmacokinetic properties, in particular clearance, in two species. To provide a rapid assessment of clearance, cassette dosing in dog was used. Modern array techniques, including the synthesis of mixtures, were used to synthesize compounds rapidly. Having selected three potential compounds as back-up candidates, they were prepared as single enantiomers and profiled in in vitro and in vivo assays and evaluated pharmacokinetically in rat and dog. These compounds are highly potent and selective HNE inhibitors, with a prolonged pharmacodynamic action. Pharmacokinetically, these compounds are comparable with 1 while they are more potent in HWB. Compound 28, however, has a higher clearance. One of these compounds, 32, was cocrystallized with HNE, and features of this structure are described and compared with the cocrystal structure of 1 in porcine pancreatic elastase. PMID:12190311

  6. Design and synthesis of pyrrolidine-5,5'-trans-lactams (5-oxo-hexahydropyrrolo[3,2-b]pyrroles) as novel mechanism-based inhibitors of human cytomegalovirus protease. 4. Antiviral activity and plasma stability.

    PubMed

    Borthwick, Alan D; Davies, Dave E; Ertl, Peter F; Exall, Anne M; Haley, Terry M; Hart, Graham J; Jackson, Deborah L; Parry, Nigel R; Patikis, Angela; Trivedi, Naimisha; Weingarten, Gordon G; Woolven, James M

    2003-10-01

    A series of chiral, (S)-proline-alpha-methylpyrrolidine-5,5-trans-lactam serine protease inhibitors has been developed as antivirals of human cytomegalovirus (HCMV). The SAR of the functionality on the proline nitrogen has shown that derivatives of para-substituted phenyl ureas > para-substituted phenyl sulfonamides > para-substituted phenyl carboxamide for activity against HCMV deltaAla protease, producing para-substituted phenyl ureas with single figure nM potency (K(i)) against the viral enzyme. The SAR of the functionality on the lactam nitrogen has defined the steric and electronic requirements for high human plasma stability while retaining good activity against HCMV protease. The combination of high potency against HCMV deltaAla protease and high human plasma stability has produced compounds with significant in vitro antiviral activity against human cytomegalovirus with the 6-hydroxymethyl benzothiazole derivative 72 being equivalent in potency to ganciclovir. The parent benzothiazole 56 had good pharmacokinetics in dogs with 29% bioavailability and good brain and ocular penetration in guinea pigs.

  7. Simultaneous Determination of Eight β-Lactam Antibiotics, Amoxicillin, Cefazolin, Cefepime, Cefotaxime, Ceftazidime, Cloxacillin, Oxacillin, and Piperacillin, in Human Plasma by Using Ultra-High-Performance Liquid Chromatography with Ultraviolet Detection.

    PubMed

    Legrand, Tiphaine; Vodovar, Dominique; Tournier, Nicolas; Khoudour, Nihel; Hulin, Anne

    2016-08-01

    A simple and rapid ultra-high-performance liquid chromatography (UHPLC) method using UV detection was developed for the simultaneous determination of eight β-lactam antibiotics in human plasma, including four penicillins, amoxicillin (AMX), cloxacillin (CLX), oxacillin (OXA), and piperacillin (PIP), and four cephalosporins, cefazolin (CFZ), cefepime (FEP), cefotaxime (CTX), and ceftazidime (CAZ). One hundred-microliter samples were spiked with thiopental as an internal standard, and proteins were precipitated by acetonitrile containing 0.1% formic acid. Separation was achieved on a pentafluorophenyl (PFP) column with a mobile phase composed of phosphoric acid (10 mM) and acetonitrile in gradient elution mode at a flow rate of 500 μl/min. Detection was performed at 230 nm for AMX, CLX, OXA, and PIP and 260 nm for CFZ, FEP, CTX, and CAZ. The total analysis time did not exceed 13 min. The method was found to be linear at concentrations ranging from 2 to 100 mg/liter for each compound, and all validation parameters fulfilled international requirements. Between- and within-run accuracy errors ranged from -5.2% to 11.4%, and precision was lower than 14.2%. This simple method requires small-volume samples and can easily be implemented in most clinical laboratories to promote the therapeutic drug monitoring of β-lactam antibiotics. The simultaneous determination of several antibiotics considerably reduces the time to results for clinicians, which may improve treatment efficiency, especially in critically ill patients. PMID:27216076

  8. β-Lactam Resistance Genes: Characterization, Epidemiology, and First Detection of blaCTX-M-1 and blaCTX-M-14 in Salmonella spp. Isolated from Poultry in Brazil-Brazil Ministry of Agriculture's Pathogen Reduction Program.

    PubMed

    Fitch, Fernanda Marques; Carmo-Rodrigues, Mirian Silva; Oliveira, Vinicius Gomes Sales; Gaspari, Marcus Vinicius; Dos Santos, Amaury; de Freitas, Josinete Barros; Pignatari, Antonio C C

    2016-03-01

    Salmonella spp. are widespread in nature; however, human infections occur mainly through ingestion of contaminated food, specially poultry and eggs. In Brazil, the Ministry of Agriculture (MAPA) oversees food production in general, with the goal of preventing transmission of pathogens through the food chain. In 2004, MAPA initiated a program to monitor and control levels of Salmonella in poultry during slaughter. This study analyzes isolates from MAPA's program for β-lactam resistance and the resistance genes involved, as well as the geographic distributions of potentially clonal populations of resistant isolates within Brazil. Initially, 1,939 Salmonella spp. isolated between 2004 and 2011 were examined. These isolates were tested for antimicrobial susceptibility, and 100 isolates resistant or intermediate to ampicillin and ceftriaxone were screened initially for the presence of blaSHV, blaTEM, blaOXA, blaPSA, blaCMY-1, and blaCMY-2 genes. There were 55 isolates whose resistance genes were not identified by this panel and these isolates are the subject of this report. These 55 isolates were differentiated into 31 distinct ribogroups, with multiple β-lactam resistance genes, including AmpC blaCMY, blaTEM, blaCTX-M-1, blaCTX-M-2, blaCTX-M-8, and blaCTX-M-14. Isolates carrying variants of blaCTX-M were identified in three geographic regions. Salmonella carrying particular genetic variants of blaCTX-M and belonging to the same ribogroup were identified from multiple poultry slaughtering facilities. In some instances, these presumptive clonal-related isolates were from facilities over 300 miles apart, indicating potential clonal spread between two geographic regions. This is the first report of blaCTX-M-1 and blaCTX-M-14 in Salmonella in Brazil.

  9. Contribution of enzymatic properties, cell permeability, and enzyme expression to microbiological activities of beta-lactams in three Bacteroides fragilis isolates that harbor a metallo-beta-lactamase gene.

    PubMed

    Rasmussen, B A; Yang, Y; Jacobus, N; Bush, K

    1994-09-01

    The metallo-beta-lactamase gene, ccrA, has been cloned from three clinical isolates of Bacteroides fragilis, TAL3636, QMCN3, and QMCN4. Although all three isolates harbored a gene encoding a potent beta-lactamase, the MICs of benzylpenicillin, piperacillin, cefotaxime, ceftazidime, imipenem, and biapenem for the three isolates varied from 4- to > 128-fold. QMCN4 was the most susceptible of the three isolates, followed by QMCN3. TAL3636 was resistant to all of the beta-lactams. Previous DNA sequence analysis of the three ccrA genes revealed that the enzymes differed at 5 amino acid residues (B. A. Rasmussen, Y. Gluzman, and F. P. Tally, Mol. Microbiol. 5:1211-1219, 1991). Biochemical characterization of the three enzymes revealed only small differences in kcat and Km values for the majority of beta-lactams tested. Thus, the 5 amino acid substitutions affected the hydrolyzing activity of the enzymes only modestly. Crypticity differences between the three isolates showed that QMCN4 was the least permeable of the isolates to cephaloridine, followed by TAL3636, and that QMCN3 was highly permeable to cephaloridine. Therefore, neither catalytic activity nor permeability was a major contributor to the dramatic differences in the MICs. Instead, microbiological susceptibility was closely related to the level of metallo-beta-lactamase present in each isolate. Both biochemical and physical studies indicated that TAL3636 produced 5- to 10-fold and 50- to 100-fold more metallo-beta-lactamase than QMCN3 and QMCN4, respectively. Therefore, the level of CcrA enzyme production is the dominant contributing factor to high-level resistance among strains harboring a ccrA gene.

  10. Metal–Arene Complexes with Indolo[3,2-c]-quinolines: Effects of Ruthenium vs Osmium and Modifications of the Lactam Unit on Intermolecular Interactions, Anticancer Activity, Cell Cycle, and Cellular Accumulation

    PubMed Central

    2013-01-01

    Six novel ruthenium(II)– and osmium(II)–arene complexes with three modified indolo[3,2-c]quinolines have been synthesized in situ starting from 2-aminoindoloquinolines and 2-pyridinecarboxaldehyde in the presence of [M(p-cymene)Cl2]2 (M = Ru, Os) in ethanol. All complexes have been characterized by elemental analysis, spectroscopic techniques (1H, 13C NMR, IR, UV–vis), and ESI mass spectrometry, while four complexes were investigated by X-ray diffraction. The complexes have been tested for antiproliferative activity in vitro in A549 (non-small cell lung), SW480 (colon), and CH1 (ovarian) human cancer cell lines and showed IC50 values between 1.3 and >80 μM. The effects of Ru vs Os and modifications of the lactam unit on intermolecular interactions, antiproliferative activity, and cell cycle are reported. One ruthenium complex and its osmium analogue have been studied for anticancer activity in vivo applied both intraperitoneally and orally against the murine colon carcinoma model CT-26. Interestingly, the osmium(II) complex displayed significant growth-inhibitory activity in contrast to its ruthenium counterpart, providing stimuli for further investigation of this class of compounds as potential antitumor drugs. PMID:23431223

  11. Macrolides and β-lactam antibiotics enhance C3b deposition on the surface of multidrug-resistant Streptococcus pneumoniae strains by a LytA autolysin-dependent mechanism.

    PubMed

    Ramos-Sevillano, Elisa; Rodríguez-Sosa, Cinthya; Díez-Martínez, Roberto; Giménez, María-José; Olmedillas, Eduardo; García, Pedro; García, Ernesto; Aguilar, Lorenzo; Yuste, Jose

    2012-11-01

    The emergence of Streptococcus pneumoniae strains displaying high levels of multidrug resistance is of great concern worldwide and a serious threat for the outcome of the infection. Modifications of the bacterial envelope by antibiotics may assist the recognition and clearance of the pathogen by the host immune system. Recognition of S. pneumoniae resistant strains by the complement component C3b was increased in the presence of specific anti-pneumococcal antibodies and subinhibitory concentrations of different macrolides and β-lactam antibiotics for all the strains investigated. However, C3b levels were unchanged in the presence of serum containing specific antibodies and sub-MICs of levofloxacin. To investigate whether LytA, the main cell wall hydrolase of S. pneumoniae, might be involved in this process, lytA-deficient mutants were constructed. In the presence of antibiotics, loss of LytA was not associated with enhanced C3b deposition on the pneumococcal surface, which confirms the importance of LytA in this interaction. The results of this study offer new insights into the development of novel therapeutic strategies using certain antibiotics by increasing the efficacy of the host immune response to efficiently recognize pneumococcal resistant strains.

  12. Lysine N[superscript zeta]-Decarboxylation Switch and Activation of the [beta]-Lactam Sensor Domain of BlaR1 Protein of Methicillin-resistant Staphylococcus aureus

    SciTech Connect

    Borbulevych, Oleg; Kumarasiri, Malika; Wilson, Brian; Llarrull1, Leticia I.; Lee, Mijoon; Hesek, Dusan; Shi, Qicun; Peng, Jeffrey; Baker, Brian M.; Mobashery, Shahriar

    2012-10-29

    The integral membrane protein BlaR1 of methicillin-resistant Staphylococcus aureus senses the presence of {beta}-lactam antibiotics in the milieu and transduces the information to the cytoplasm, where the biochemical events that unleash induction of antibiotic resistance mechanisms take place. We report herein by two-dimensional and three-dimensional NMR experiments of the sensor domain of BlaR1 in solution and by determination of an x-ray structure for the apo protein that Lys-392 of the antibiotic-binding site is posttranslationally modified by N{sup {zeta}}-carboxylation. Additional crystallographic and NMR data reveal that on acylation of Ser-389 by antibiotics, Lys-392 experiences N{sup {zeta}}-decarboxylation. This unique process, termed the lysine N{sup {zeta}}-decarboxylation switch, arrests the sensor domain in the activated ('on') state, necessary for signal transduction and all the subsequent biochemical processes. We present structural information on how this receptor activation process takes place, imparting longevity to the antibiotic-receptor complex that is needed for the induction of the antibiotic-resistant phenotype in methicillin-resistant S. aureus.

  13. Macrolides and β-Lactam Antibiotics Enhance C3b Deposition on the Surface of Multidrug-Resistant Streptococcus pneumoniae Strains by a LytA Autolysin-Dependent Mechanism

    PubMed Central

    Ramos-Sevillano, Elisa; Rodríguez-Sosa, Cinthya; Díez-Martínez, Roberto; Giménez, María-José; Olmedillas, Eduardo; García, Pedro; García, Ernesto; Aguilar, Lorenzo

    2012-01-01

    The emergence of Streptococcus pneumoniae strains displaying high levels of multidrug resistance is of great concern worldwide and a serious threat for the outcome of the infection. Modifications of the bacterial envelope by antibiotics may assist the recognition and clearance of the pathogen by the host immune system. Recognition of S. pneumoniae resistant strains by the complement component C3b was increased in the presence of specific anti-pneumococcal antibodies and subinhibitory concentrations of different macrolides and β-lactam antibiotics for all the strains investigated. However, C3b levels were unchanged in the presence of serum containing specific antibodies and sub-MICs of levofloxacin. To investigate whether LytA, the main cell wall hydrolase of S. pneumoniae, might be involved in this process, lytA-deficient mutants were constructed. In the presence of antibiotics, loss of LytA was not associated with enhanced C3b deposition on the pneumococcal surface, which confirms the importance of LytA in this interaction. The results of this study offer new insights into the development of novel therapeutic strategies using certain antibiotics by increasing the efficacy of the host immune response to efficiently recognize pneumococcal resistant strains. PMID:22890762

  14. Evaluation of the in vitro activity of six broad-spectrum beta-lactam antimicrobial agents tested against over 2,000 clinical isolates from 22 medical centers in Japan. Japan Antimicrobial Resistance Study Group.

    PubMed

    Yamaguchi, K; Mathai, D; Biedenbach, D J; Lewis, M T; Gales, A C; Jones, R N

    1999-06-01

    Numerous broad-spectrum beta-lactam antimicrobial agents have been introduced into medical practice since 1985. Although several of these compounds have advanced, infectious disease therapy resistances to them has also emerged world-wide. In 1997, a Japanese 22 medical center investigation was initiated to assess the continued utility of these agents (oxacillin or piperacillin, ceftazidime, cefepime, cefpirome, cefoperazone/sulbactam [C/S], imipenem). The participating medical centers represented a wide geographic distribution, and a common protocol and reagents were applied. Three control strains and a set of challenge organisms were provided to participant centers. Etest (AB BIODISK, Solna, Sweden) strips were used in concurrent tests of these organisms and a qualitative determination of participant skills in the identification of resistant and susceptible phenotypes was established. The quantitative controls demonstrated 97.7-99.2% of MIC values within established QC limits, and the qualitative (susceptibility category) controls documented a 97.3% agreement of participant results with that of reference values (1,320 total results). Only 0.2% of values were false-susceptible errors. After the participant quality was assured, a total of 2,015 clinical strains were tested (10 strains from 10 different organism groups including methicillin-susceptible Staphylococcus aureus and coagulase-negative staphylococci [CoNS], Escherichia coli, Klebsiella spp., Citrobacter freundii, Enterobacter spp., indole-positive Proteae, Serratia spp., Acinetobacter spp., and Pseudomonas aeruginosa). The staphylococci were uniformly susceptible to all drugs tested except ceftazidime (MIC90, 24 micrograms/ml) that had a potency six- to 12-fold less than either cefepime or cefpirome. Only 3.7 and 45.1% of S. aureus and CoNS were susceptible to ceftazidime, respectively. Among E. coli and Klebsiella spp. the rank order of antimicrobial spectrum was imipenem = "fourth

  15. In Vitro Activities of the β-Lactamase Inhibitors Clavulanic Acid, Sulbactam, and Tazobactam Alone or in Combination with β-Lactams against Epidemiologically Characterized Multidrug-Resistant Acinetobacter baumannii Strains

    PubMed Central

    Higgins, Paul G.; Wisplinghoff, Hilmar; Stefanik, Danuta; Seifert, Harald

    2004-01-01

    Acinetobacter baumannii is an important nosocomial pathogen usually in the context of serious underlying disease. Multidrug resistance in these organisms is frequent. The β-lactamase inhibitors clavulanic acid, sulbactam, and tazobactam have intrinsic activity against Acinetobacter strains. To evaluate their potential therapeutic usefulness, we determined the in vitro activity of ampicillin, sulbactam, ampicillin-sulbactam, cefoperazone, cefoperazone-sulbactam, piperacillin, piperacillin-sulbactam, tazobactam, piperacillin-tazobactam, amoxicillin, clavulanic acid, amoxicillin-clavulanic acid, ticarcillin, and ticarcillin-clavulanic acid against multidrug-resistant A. baumannii. All isolates were epidemiologically characterized by RAPD [random(ly) amplified polymorphic DNA] analysis and/or pulsed-field gel electrophoresis and represented different strain types, including sporadic strains, as well as outbreak-related strains. The MICs were determined by agar dilution on Mueller-Hinton agar (using fixed concentrations, as well as fixed ratios for β-lactamase inhibitors) and the E-test. The majority of E-test results were within two dilutions of those recorded by agar dilution, with the exception of piperacillin-tazobactam. Sulbactam was superior to clavulanic acid and tazobactam and may represent an alternative treatment option for infections due to multiresistant A. baumannii strains. β-Lactamase inhibitors have intrinsic activity but do not enhance activity of β-lactams against A. baumannii. Testing with the inhibitor added at a fixed concentration as recommended for piperacillin-tazobactam and ticarcillin-clavulanic acid by the National Committee for Clinical Laboratory Standards may falsely suggest high activity or gives uninterpretable results due to trailing. If combinations are used for testing, fixed ratios may give more useful results. PMID:15105109

  16. Selective spectrophotometric determination of phenolic beta-lactam antibiotics.

    PubMed

    Salem, Hesham; Saleh, Gamal A

    2002-06-15

    Two simple and selective spectrophotometric methods were developed for the quantitative determination of cefoperazone sodium, cefadroxil monohydrate, cefprozil anhydrous and amoxicillin trihydrate in pure forms as well as in their pharmaceutical formulations. The methods are based on the selective oxidation of these drugs with either Ce (IV) or Fe (III) in acid medium to give an intense yellow coloured product (lambda(max)=397 nm). The reaction conditions were studied and optimized. Beer's plots were obeyed in a general concentration range of 5-30 microg ml(-1) with correlation coefficients not less than 0.9979 for the four drugs with the two reagents. The methods are successfully applied to the analysis of pharmaceutical formulations containing amoxicillin, either alone or in combination with potassium clavulanate, flucloxacillin or dicloxacillin. They were also applied to the analysis of the other three studied drugs in vials, capsules, tablets and suspensions with good recovery; percent ranged from 99.7 (+/-0.46) to 100.32 (+/-1.05) in the Ce (IV) method and 99.6 (+/-0.50) to 100.3 (+/-1.32) in the Fe (III) method. Interferences from other antibiotics and additives products were investigated.

  17. Beckmann rearrangement of ketoximes to lactams by triphosphazene catalyst.

    PubMed

    Hashimoto, Masaharu; Obora, Yasushi; Sakaguchi, Satoshi; Ishii, Yasutaka

    2008-04-01

    Triphosphazene, 1,3,5-triazo-2,4,6-triphosphorine-2,2,4,4,6,6-chloride (TAPC), was found to be an efficient catalyst for the Beckmann rearrangement of cyclohexanone oxime and cyclododecanone oxime to epsilon-caprolactam and laurolactam, which are raw materials of nylon-6 and nylon-12, respectively.

  18. Vicenistatin, a novel 20-membered macrocyclic lactam antitumor antibiotic.

    PubMed

    Shindo, K; Kamishohara, M; Odagawa, A; Matsuoka, M; Kawai, H

    1993-07-01

    A new antitumor antibiotic vicenistatin was isolated from the culture broth of Streptomyces sp. HC34. The structure of vicenistatin was elucidated by NMR spectral analysis. Vicenistatin exhibited antitumor activity against human colon carcinoma Co-3 in the xenograft model. PMID:8360102

  19. Industrial enzymatic production of cephalosporin-based beta-lactams.

    PubMed

    Barber, Michael S; Giesecke, Ulrich; Reichert, Arno; Minas, Wolfgang

    2004-01-01

    Cephalosporins are chemically closely related to penicillins both work by inhibiting the cell wall synthesis of bacteria. The first generation cephalosporins entered the market in 1964. Second and third generation cephalosporins were subsequently developed that were more powerful than the original products. Fourth generation cephalosporins are now reaching the market. Each newer generation of cephalosporins has greater Gram-negative antimicrobial properties than the preceding generation. Conversely, the 'older' generations of cephalosporins have greater Gram-positive (Staphylococcus and Streptococcus) coverage than the 'newer' generations. Frequency of dosing decreases and palatability generally improve with increasing generations. The advent of fourth generation cephalosporins with the launch of cefepime extended the spectrum against Gram-positive organisms without a significant loss of activity towards Gram-negative bacteria. Its greater stability to beta-lactamases increases its efficacy against drug-resistant bacteria. In this review we present the current situation of this mature market. In addition, we present the current state of the technologies employed for the production of cephalosporins, focusing on the new and environmentally safer 'green' routes to the products. Starting with the fermentation and purification of CPC, enzymatic conversion in conjunction with aqueous chemistry will lead to some key intermediates such as 7-ACA, TDA and TTA, which then can be converted into the active pharmaceutical ingredient (API), again applying biocatalytic technologies and aqueous chemistry. Examples for the costing of selected products are provided as well. PMID:15719556

  20. Evaluation of melanotan-II, a superpotent cyclic melanotropic peptide in a pilot phase-I clinical study.

    PubMed

    Dorr, R T; Lines, R; Levine, N; Brooks, C; Xiang, L; Hruby, V J; Hadley, M E

    1996-01-01

    A pilot phase I study was conducted with a cyclic heptapeptide analog of alpha-melanocyte stimulating hormone (alpha-MSH). The lactam-bridged molecule, called Melanotan-II (MT-II), has the structure Ac-Nle4-Asp5-His6-D-Phe7-Arg8-Trp9-Lys10 alpha-MSH4-10-NH2 (MT-II) and has superpotent melanotropic activity in vitro. A single-blind, alternating day (saline or MT-II), placebo-controlled trial was conducted in 3 normal male volunteers at the starting dose of 0.01 mg/kg of MT-II. Subcutaneous injections of MT-II or saline were given daily (Monday-Friday) for 2 consecutive weeks. Two subjects were escalated by 0.005 mg/kg increments to 0.03 mg/kg and one to 0.025 mg/kg. The 0.03 mg/kg dose produced Grade II somnolence and fatigue in one of two subjects (WHO standards). Mild nausea, not requiring antiemetic treatment, was reported at most MT-II dose levels. A stretching and yawning complex appeared to correlate with the onset of spontaneous, penile erections which were intermittently experienced for 1-5 hours after MT-II dosing, depending on the MT-II dose. Two subjects had increased pigmentation in the face, upper body and buttock, as measured by quantitative reflectance and by visual perception 1 week after MT-II dosing ended. These results demonstrate that MT-II has tanning activity in humans given only 5 low doses every other day by subcutaneous injection. The recommended single MT-II dose for future Phase I studies is 0.025 mg/kg/day.

  1. Binding of faropenem and other beta-lactam agents to penicillin-binding proteins of pneumococci with various beta-lactam susceptibilities.

    PubMed

    Kosowska-Shick, Klaudia; McGhee, Pamela; Appelbaum, Peter C

    2009-05-01

    Faropenem demonstrated low MICs (< or = 1 microg/ml) for all penicillin-susceptible and nonsusceptible pneumococci and exhibited very strong abilities to bind to Streptococcus pneumoniae penicillin-binding proteins (PBPs), except for PBP2X. The lower faropenem affinity for PBP2X did not affect MICs for any strains tested, and only imipenem had lower MICs, with much lower binding affinities for all PBPs tested, than faropenem. PMID:19237649

  2. Binding of Faropenem and Other β-Lactam Agents to Penicillin-Binding Proteins of Pneumococci with Various β-Lactam Susceptibilities▿

    PubMed Central

    Kosowska-Shick, Klaudia; McGhee, Pamela; Appelbaum, Peter C.

    2009-01-01

    Faropenem demonstrated low MICs (≤1 μg/ml) for all penicillin-susceptible and nonsusceptible pneumococci and exhibited very strong abilities to bind to Streptococcus pneumoniae penicillin-binding proteins (PBPs), except for PBP2X. The lower faropenem affinity for PBP2X did not affect MICs for any strains tested, and only imipenem had lower MICs, with much lower binding affinities for all PBPs tested, than faropenem. PMID:19237649

  3. Theoretical study of the alkaline hydrolysis of an aza-β-lactam derivative of clavulanic acid

    NASA Astrophysics Data System (ADS)

    Garcías, Rafael C.; Coll, Miguel; Donoso, Josefa; Muñoz, Francisco

    2003-04-01

    DFT calculations based on the hybrid functional B3LYP/6-31+G * were used to study the alkaline hydrolysis of an aza-clavulanic acid, which results from the substitution of the carbon atom at position 6 in clavulanic acid by a nitrogen atom. The presence of the nitrogen atom endows the compound with special properties; in fact, once formed, the tetrahedral intermediate can evolve with cleavage of the N 4-C 7 or N 6-C 7 bond, which obviously leads to different reaction products. These differential bond cleavages may play a central role in the inactivation of β-lactamases, so the compound may be a powerful inactivator of these enzymes.

  4. Penicillin-bound polyacrylate nanoparticles: restoring the activity of beta-lactam antibiotics against MRSA.

    PubMed

    Turos, Edward; Reddy, G Suresh Kumar; Greenhalgh, Kerriann; Ramaraju, Praveen; Abeylath, Sampath C; Jang, Seyoung; Dickey, Sonja; Lim, Daniel V

    2007-06-15

    This report describes the preparation of antibacterially active emulsified polyacrylate nanoparticles in which a penicillin antibiotic is covalently conjugated onto the polymeric framework. These nanoparticles were prepared in water by emulsion polymerization of an acrylated penicillin analogue pre-dissolved in a 7:3 (w:w) mixture of butyl acrylate and styrene in the presence of sodium dodecyl sulfate (surfactant) and potassium persulfate (radical initiator). Dynamic light scattering analysis and atomic force microscopy images show that the emulsions contain nanoparticles of approximately 40 nm in diameter. The nanoparticles have equipotent in vitro antibacterial properties against methicillin-susceptible and methicillin-resistant forms of Staphylococcus aureus and indefinite stability toward beta-lactamase. PMID:17420125

  5. Potent combinations of beta-lactam antibiotics using the beta-lactamase inhibition principle.

    PubMed

    Greenwood, D; O'Grady, F

    1975-01-01

    Several penicillins known to be stable to enterobacterial beta-lactamases were tested in combination with beta-lactamase-sensitive penicillins and cephalosporins in a turbidimetric system. Nafcillin was found to be the best beta-lactamase inhibitor amongst agents presently available commercially, but the related, new semi-synthetic penicillin BRL 1437 (2-isopropoxy-1-naphthylpenicillin) was consistently found to be superior. Using 103 ampicillin-resistant coliform bacilli and antibiotic levels achievable in urine, cephalothin or cephaloridine alone achieved long-term suppression of growth (greater than 20 h) of 16 and 13% of strains, respectively, while the additional presence of BRL 1437 suppressed growth for longer than 20 h of 81% of the remaining strains. Even where 'success' was not achieved according to these stringent criteria, regrowth was significantly delayed by the presence of BRL 1437. Suppression of growth for longer than 20 h by BRL 1437 plus cephalothin was achieved with all of the 46 Escherichia coli strains tested. Antibiotic combinations were also studied in an in vitro model which stimulates the hydrokinetic features of the urinary bladder. Suppression of the growth of two highly resistant E. coli strains was achieved in this system, for therapeutically acceptable periods of time, with combinations of cephalothin or cephaloridine with BRL 1437, but not nafcillin.

  6. Beta-lactam susceptibility of coagulase-negative staphylococci causing catheter sepsis in pediatric patients.

    PubMed

    Marcon, M J; Nahata, M C; Powell, D A; Lisby-Sutch, S M

    1990-01-01

    We evaluated standard oxacillin and methicillin disk diffusion (DD) and broth microdilution (MD)-MIC tests with and without 2% NaCl for detecting heteroresistance among 47 blood isolates of coagulase-negative staphylococci (CNS) causing catheter sepsis in pediatric patients. The 24-hr oxacillin DD test detected the greatest number (40) of apparent hetero-resistant isolates, but methicillin DD and oxacillin MD-MIC with 2% NaCl performed equally as well (38 and 37 resistant isolates, respectively). An additional 24-hr incubation did not significantly increase the number of apparent heteroresistant isolates detected by these methods. Discrepant results with the various test methods occurred most commonly among Staphylococcus epidermidis isolates with MD-MIC values near the breakpoint concentrations for interpretation of susceptible and resistant strains. For detection of heteroresistance among the CNS, we encourage use of standard oxacillin DD and MD-MIC tests but would suggest that isolates with MIC values ranging from 1-2 micrograms/ml be interpreted cautiously until clinical studies demonstrate the efficacy of treating patients with infections caused by such strains.

  7. Benzoxazinoids-cyclic hydroxamic acids, lactams and their corresponding glucosides in the genus Aphelandra (Acanthaceae).

    PubMed

    Baumeler, A; Hesse, M; Werner, C

    2000-01-01

    An improved method of sample preparation and simultaneous HPLC separation was developed that allowed the separation of 2,4-dihydroxy-1,4-benzoxazine-3(4H)-one (DIBOA), 2,4-dihydroxy-7-methoxy-1,4-benzoxazine-3(4H)-one (DIMBOA), 2-hydroxy-1,4-benzoxazine-3(2H)-one (HBOA), 2-hydroxy-7-methoxy-1,4-benzoxazine-3(2H)-one (HMBOA) and their corresponding glucosides as well as the benzoxazolinones BOA and MBOA. The amount and distribution of these compounds was determined in the roots of Aphelandra squarrosa and A. fuscopunctata plants. There is a significant difference in the amount and distribution of this substance class in the two species analyzed. The results are discussed in relation to their function as defence compounds and allelochemicals.

  8. Asymmetric Reduction of Lactam-Based β-Aminoacrylates. Synthesis of Heterocyclic β(2)-Amino Acids.

    PubMed

    Campello, Hugo Rego; Parker, Jeremy; Perry, Matthew; Ryberg, Per; Gallagher, Timothy

    2016-08-19

    The ability to affect asymmetric reduction of heterocyclic β-aminoacrylates 1 (n = 1-3) has been assessed with pyrrolidine and piperidone variants generating the corresponding N-heterocyclic β(2)-amino acids 3b and 5b with high enantioselectivity (≥97% ee) using a Rh/WALPHOS catalyst combination. The use of the carboxylic acid substrate was essential; the corresponding esters do undergo reduction but led to racemic products. The seven-ring azepanone variant (as the carboxylic acid 9b) underwent reduction, but only a minimal level of asymmetric induction was observed. PMID:27508307

  9. Process design and evaluation of production of bioethanol and β-lactam antibiotic from lignocellulosic biomass.

    PubMed

    Kim, Sung Bong; Park, Chulhwan; Kim, Seung Wook

    2014-11-01

    To design biorefinery processes producing bioethanol from lignocellulosic biomass with dilute acid pretreatment, biorefinery processes were simulated using the SuperPro Designer program. To improve the efficiency of biomass use and the economics of biorefinery, additional pretreatment processes were designed and evaluated, in which a combined process of dilute acid and aqueous ammonia pretreatments, and a process of waste media containing xylose were used, for the production of 7-aminocephalosporanic acid. Finally, the productivity and economics of the designed processes were compared.

  10. Conformation of eight-membered benzoannulated lactams by combined NMR and DFT studies.

    PubMed

    Witosińska, Agnieszka; Musielak, Bogdan; Serda, Paweł; Owińska, Maria; Rys, Barbara

    2012-11-01

    The title compounds were synthesized, and their structure and conformational behavior in solution (NMR and DFT), in the gas phase (DFT), and, for some of them, in the solid state (X-ray) were investigated. The variable-temperature NMR spectra were employed to determine the conformational equilibria and the activation energy of the conformational changes of the eight-membered ring. The coalescence effects are assigned to racemization of the chiral ground-state conformation with a ring inversion barrier in the range of 38-100 kJ mol(-1) depending on the relative setting of the two strong conformational constraints: benzoannulation and the amide function. The second conformational process, interconversion between two different conformers, in the molecules of benzo[c]azocin-3-one, benzo[d]azocin-2-one, and benzo[d]azocin-4-one was observed. The natures of the conformers observed in solution were elucidated by analysis of experimental and calculated NMR data. The present results are discussed in conjunction with previous experimental and theoretical data on (Z,Z)-cyclooctadienes and their benzo analogues.

  11. Clavulanic acid: a beta-lactamase-inhiting beta-lactam from Streptomyces clavuligerus.

    PubMed

    Reading, C; Cole, M

    1977-05-01

    A novel beta-lactamase inhibitor has been isolated from Streptomyces clavuligerus ATCC 27064 and given the name clavulanic acid. Conditions for the cultivation of the organism and detection and isolation of clavulanic acid are described. This compound resembles the nucleus of a penicillin but differs in having no acylamino side chain, having oxygen instead of sulfur, and containing a beta-hydroxyethylidine substituent in the oxazolidine ring. Clavulanic acid is a potent inhibitor of many beta-lactamases, including those found in Escherichia coli (plasmid mediated), Klebsiella aerogenes, Proteus mirabilis, and Staphylococcus aureus, the inhibition being of a progressive type. The cephalosporinase type of beta-lactamase found in Pseudomonas aeruginosa and Enterobacter cloacae P99 and the chromosomally mediated beta-lactamase of E. coli are less well inhibited. The minimum inhibitory concentrations of ampicillin and cephaloridine against beta-lactamase-producing, penicillin-resistant strains of S. aureus, K. aerogenes, P. mirabilis, and E. coli have been shown to be considerably reduced by the addition of low concentrations of clavulanic acid.

  12. An efficient system for intracellular delivery of beta-lactam antibiotics to overcome bacterial resistance.

    PubMed

    Abed, Nadia; Saïd-Hassane, Fatouma; Zouhiri, Fatima; Mougin, Julie; Nicolas, Valérie; Desmaële, Didier; Gref, Ruxandra; Couvreur, Patrick

    2015-01-01

    The "Golden era" of antibiotics is definitely an old story and this is especially true for intracellular bacterial infections. The poor intracellular bioavailability of antibiotics reduces the efficency of many treatments and thereby promotes resistances. Therefore, the development of nanodevices coupled with antibiotics that are capable of targeting and releasing the drug into the infected-cells appears to be a promising solution to circumvent these complications. Here, we took advantage of two natural terpenes (farnesyl and geranyl) to design nanodevices for an efficient intracellular delivery of penicillin G. The covalent linkage between the terpene moieties and the antibiotic leads to formation of prodrugs that self-assemble to form nanoparticles with a high drug payload between 55-63%. Futhermore, the addition of an environmentally-sensitive bond between the antibiotic and the terpene led to an efficient antibacterial activity against the intracellular pathogen Staphylococcus aureus with reduced intracellular replication of about 99.9% compared to untreated infected cells. Using HPLC analysis, we demonstrated and quantified the intracellular release of PenG when this sensitive-bond (SB) was present on the prodrug, showing the success of this technology to deliver antibiotics directly into cells.

  13. [Evaluation and characterization of 125 patients with a history of reaction to beta-lactams].

    PubMed

    Beltrán-Sierra, Kaddy Juliana; Sánchez, Jorge; Cardona, Ricardo

    2016-01-01

    Antecedentes: Las reacciones a betalactámicos son frecuentes y es difícil establecer la relación entre el medicamento y los síntomas. Objetivo: Describir las características clínicas y explorar los mecanismos inmunológicos de pacientes con sospecha de reacción adversa a betalactámicos. Métodos: Estudio retrospectivo de pacientes con antecedentes de reacción a betalactámicos y pruebas para reacción con medicamentos. Resultados: De 125 pacientes, 71 eran mujeres (56.8%); 73 tenían historia de reacción inmediata y 52 de reacción tardía; Se realizaron 590 pruebas de alergia: medición de IgE específica, pruebas intradérmicas, de parche y de provocación. Los medicamentos más relacionados fueron amoxicilina, en 62 pacientes (49.6%), penicilina cristalina en 17 (13.6%), penicilina benzatínica en 15 (12%) y cefalexina en 13 (10.4%). La severidad de la reacción fue leve en la mayoría (82%). 7.7% tuvo antecedente de anafilaxia y 10.8% consultaba por una prueba cutánea con penicilina positiva, sin antecedente de reacción. Solo 6.7% resultó con una prueba positiva. Más de 62% inició los estudios dos años después de la reacción por la que consultó. Conclusiones: La historia clínica es insuficiente para determinar la alergia a betalactámicos. En nuestra muestra, pocos pacientes con antecedente de reacción a betalactámicos tuvo evidencia de reacciones inmunológicamente mediadas.

  14. Potent combinations of beta-lactam antibiotics using the beta-lactamase inhibition principle.

    PubMed

    Greenwood, D; O'Grady, F

    1975-01-01

    Several penicillins known to be stable to enterobacterial beta-lactamases were tested in combination with beta-lactamase-sensitive penicillins and cephalosporins in a turbidimetric system. Nafcillin was found to be the best beta-lactamase inhibitor amongst agents presently available commercially, but the related, new semi-synthetic penicillin BRL 1437 (2-isopropoxy-1-naphthylpenicillin) was consistently found to be superior. Using 103 ampicillin-resistant coliform bacilli and antibiotic levels achievable in urine, cephalothin or cephaloridine alone achieved long-term suppression of growth (greater than 20 h) of 16 and 13% of strains, respectively, while the additional presence of BRL 1437 suppressed growth for longer than 20 h of 81% of the remaining strains. Even where 'success' was not achieved according to these stringent criteria, regrowth was significantly delayed by the presence of BRL 1437. Suppression of growth for longer than 20 h by BRL 1437 plus cephalothin was achieved with all of the 46 Escherichia coli strains tested. Antibiotic combinations were also studied in an in vitro model which stimulates the hydrokinetic features of the urinary bladder. Suppression of the growth of two highly resistant E. coli strains was achieved in this system, for therapeutically acceptable periods of time, with combinations of cephalothin or cephaloridine with BRL 1437, but not nafcillin. PMID:1102266

  15. Susceptibility to β-lactams and quinolones of Enterobacteriaceae isolated from urinary tract infections in outpatients

    PubMed Central

    Marchisio, Martín; Porto, Ayelén; Joris, Romina; Rico, Marina; Baroni, María R.; Di Conza, José

    2015-01-01

    Abstract The antibiotic susceptibility profile was evaluated in 71 Enterobacteriaceae isolates obtained from outpatient urine cultures in July 2010 from two health institutions in Santa Fe, Argentina. The highest rates of antibiotic resistance were observed for ampicillin (AMP) (69%), trimethoprim/sulfamethoxazole (TMS) (33%), and ciprofloxacin (CIP) (25%). Meanwhile, 21% of the isolates were resistant to three or more tested antibiotics families. Thirty integron-containing bacteria (42.3%) were detected, and a strong association with TMS resistance was found. Third generation cephalosporin resistance was detected in only one Escherichia coli isolate, and it was characterized as a bla CMY-2 carrier. No plasmid-mediated quinolone resistance (PMQR) was found. Resistance to fluoroquinolone in the isolates was due to alterations in QRDR regions. Two mutations in GyrA (S83L, D87N) and one in ParC (S80I) were observed in all CIP-resistant E. coli. It was determined to be the main phylogenetic groups in E. coli isolates. Minimum Inhibitory Concentration (MIC) values against nalidixic acid (NAL), levofloxacin (LEV), and CIP were determined for 63 uropathogenic E. coli isolates as MIC50 of 4 μg/mL, 0.03125 μg/mL, and 0.03125 μg/mL, respectively, while the MIC90 values of the antibiotics were determined as 1024 μg/mL, 64 μg/mL, and 16 μg/mL, respectively. An association between the phylogenetic groups, A and B1 with fluoroquinolone resistance was observed. These results point to the importance of awareness of the potential risk associated with empirical treatment with both the families of antibiotics. PMID:26691475

  16. Beta-lactam susceptibility of coagulase-negative staphylococci causing catheter sepsis in pediatric patients.

    PubMed

    Marcon, M J; Nahata, M C; Powell, D A; Lisby-Sutch, S M

    1990-01-01

    We evaluated standard oxacillin and methicillin disk diffusion (DD) and broth microdilution (MD)-MIC tests with and without 2% NaCl for detecting heteroresistance among 47 blood isolates of coagulase-negative staphylococci (CNS) causing catheter sepsis in pediatric patients. The 24-hr oxacillin DD test detected the greatest number (40) of apparent hetero-resistant isolates, but methicillin DD and oxacillin MD-MIC with 2% NaCl performed equally as well (38 and 37 resistant isolates, respectively). An additional 24-hr incubation did not significantly increase the number of apparent heteroresistant isolates detected by these methods. Discrepant results with the various test methods occurred most commonly among Staphylococcus epidermidis isolates with MD-MIC values near the breakpoint concentrations for interpretation of susceptible and resistant strains. For detection of heteroresistance among the CNS, we encourage use of standard oxacillin DD and MD-MIC tests but would suggest that isolates with MIC values ranging from 1-2 micrograms/ml be interpreted cautiously until clinical studies demonstrate the efficacy of treating patients with infections caused by such strains. PMID:2331855

  17. An efficient system for intracellular delivery of beta-lactam antibiotics to overcome bacterial resistance

    PubMed Central

    Abed, Nadia; Saïd-Hassane, Fatouma; Zouhiri, Fatima; Mougin, Julie; Nicolas, Valérie; Desmaële, Didier; Gref, Ruxandra; Couvreur, Patrick

    2015-01-01

    The “Golden era” of antibiotics is definitely an old story and this is especially true for intracellular bacterial infections. The poor intracellular bioavailability of antibiotics reduces the efficency of many treatments and thereby promotes resistances. Therefore, the development of nanodevices coupled with antibiotics that are capable of targeting and releasing the drug into the infected-cells appears to be a promising solution to circumvent these complications. Here, we took advantage of two natural terpenes (farnesyl and geranyl) to design nanodevices for an efficient intracellular delivery of penicillin G. The covalent linkage between the terpene moieties and the antibiotic leads to formation of prodrugs that self-assemble to form nanoparticles with a high drug payload between 55–63%. Futhermore, the addition of an environmentally-sensitive bond between the antibiotic and the terpene led to an efficient antibacterial activity against the intracellular pathogen Staphylococcus aureus with reduced intracellular replication of about 99.9% compared to untreated infected cells. Using HPLC analysis, we demonstrated and quantified the intracellular release of PenG when this sensitive-bond (SB) was present on the prodrug, showing the success of this technology to deliver antibiotics directly into cells. PMID:26311631

  18. Synergistic antibacterial effects of β-lactam antibiotic combined with silver nanoparticles

    NASA Astrophysics Data System (ADS)

    Li, Ping; Li, Juan; Wu, Changzhu; Wu, Qingsheng; Li, Jian

    2005-09-01

    The bactericidal action of silver (0) nanoparticles and amoxicillin on Escherichia coli is studied, respectively. Increasing concentration of both amoxicillin (0-0.525 mg ml-1) and silver nanoparticles (0-40 µg ml-1) showed a higher antibacterial effect in Luria-Bertani (LB) medium. Escherichia coli cells have different bactericidal sensitivity to them. When amoxicillin and silver nanoparticles are combined, it results in greater bactericidal efficiency on Escherichia coli cells than when they were applied separately. Dynamic tests on bacterial growth indicated that exponential and stationary phases are greatly decreased and delayed in the synergistic effect of amoxicillin and silver nanoparticles. In addition, the effect induced by a preincubation with silver nanoparticles is examined. The results show that solutions with more silver nanoparticles have better antimicrobial effects. One hypothesized mechanism is proposed to explain this phenomenon.

  19. Replicon typing of plasmids encoding resistance to newer beta-lactams.

    PubMed

    Carattoli, Alessandra; Miriagou, Vivi; Bertini, Alessia; Loli, Alexandra; Colinon, Celine; Villa, Laura; Whichard, Jean M; Rossolini, Gian Maria

    2006-07-01

    Polymerase chain reaction-based replicon typing represents a novel method to describe the dissemination and follow the evolution of resistance plasmids. We used this approach to study 26 epidemiologically unrelated Enterobacteriaceae and demonstrate the dominance of incompatibility (Inc) A/C or Inc N-related plasmids carrying some emerging resistance determinants to extended-spectrum cephalosporins and carbapenems. PMID:16836838

  20. Impact of the New Delhi metallo-beta-lactamase on beta-lactam antibiotics

    PubMed Central

    Zmarlicka, Monika T; Nailor, Michael D; Nicolau, David P

    2015-01-01

    Since the first New Delhi metallo-beta-lactamase (NDM) report in 2009, NDM has spread globally causing various types of infections. NDM-positive organisms produce in vitro resistance phenotypes to carbapenems and many other antimicrobials. It is thus surprising that the literature examining clinical experiences with NDM does not report corresponding poor clinical outcomes. There are many instances where good clinical outcomes are described, despite a mismatch between administered antimicrobials and resistant in vitro susceptibilities. Available in vitro data for either monotherapy or combination therapy does not provide an explanation for these observations. However, animal studies do begin to shed more light on this phenomenon. They imply that the in vivo expression of NDM may not confer clinical resistance to all cephalosporin and carbapenem antibiotics as predicted by in vitro testing but other resistance mechanisms need to be present to generate a resistant phenotype. As such, previously abandoned therapies, particularly carbapenems and beta-lactamase inhibitor combinations, may retain utility against infections caused by NDM producers. PMID:26345624

  1. Semisynthetic beta-lactam antibiotics. II Cephalosporin derivatives in the naphthalene series. Chemical and microbiological properties.

    PubMed

    Quaresima, E; Tinti, M O; Foresta, P; De Witt, P; Ramacci, M T

    1979-12-01

    A seris of new 7-acylamidocephalosporins, containing a substituted naphthalene moiety in the side chain, has been prepared and tested for their in vitro antibacterial activity. Some observations are made on the structure-activity relationships.

  2. Replicon typing of plasmids encoding resistance to newer beta-lactams.

    PubMed

    Carattoli, Alessandra; Miriagou, Vivi; Bertini, Alessia; Loli, Alexandra; Colinon, Celine; Villa, Laura; Whichard, Jean M; Rossolini, Gian Maria

    2006-07-01

    Polymerase chain reaction-based replicon typing represents a novel method to describe the dissemination and follow the evolution of resistance plasmids. We used this approach to study 26 epidemiologically unrelated Enterobacteriaceae and demonstrate the dominance of incompatibility (Inc) A/C or Inc N-related plasmids carrying some emerging resistance determinants to extended-spectrum cephalosporins and carbapenems.

  3. Replicon Typing of Plasmids Encoding Resistance to Newer β-Lactams

    PubMed Central

    Miriagou, Vivi; Bertini, Alessia; Loli, Alexandra; Colinon, Celine; Villa, Laura; Whichard, Jean M.; Rossolini, Gian Maria

    2006-01-01

    Polymerase chain reaction–based replicon typing represents a novel method to describe the dissemination and follow the evolution of resistance plasmids. We used this approach to study 26 epidemiologically unrelated Enterobacteriaceae and demonstrate the dominance of incompatibility (Inc) A/C or Inc N-related plasmids carrying some emerging resistance determinants to extended-spectrum cephalosporins and carbapenems. PMID:16836838

  4. Clavulanic Acid: a Beta-Lactamase-Inhibiting Beta-Lactam from Streptomyces clavuligerus

    PubMed Central

    Reading, C.; Cole, M.

    1977-01-01

    A novel β-lactamase inhibitor has been isolated from Streptomyces clavuligerus ATCC 27064 and given the name clavulanic acid. Conditions for the cultivation of the organism and detection and isolation of clavulanic acid are described. This compound resembles the nucleus of a penicillin but differs in having no acylamino side chain, having oxygen instead of sulfur, and containing a β-hydroxyethylidine substituent in the oxazolidine ring. Clavulanic acid is a potent inhibitor of many β-lactamases, including those found in Escherichia coli (plasmid mediated), Klebsiella aerogenes, Proteus mirabilis, and Staphylococcus aureus, the inhibition being of a progressive type. The cephalosporinase type of β-lactamase found in Pseudomonas aeruginosa and Enterobacter cloacae P99 and the chromosomally mediated β-lactamase of E. coli are less well inhibited. The minimum inhibitory concentrations of ampicillin and cephaloridine against β-lactamase-producing, penicillin-resistant strains of S. aureus, K. aerogenes, P. mirabilis, and E. coli have been shown to be considerably reduced by the addition of low concentrations of clavulanic acid. Images PMID:879738

  5. Constitutively vancomycin-resistant Enterococcus faecium resistant to synergistic beta-lactam combinations.

    PubMed Central

    Green, M; Binczewski, B; Pasculle, A W; Edmund, M; Barbadora, K; Kusne, S; Shlaes, D M

    1993-01-01

    Vancomycin resistance among enterococci has recently been recognized. Synergy between vancomycin and penicillin has been shown in vitro for isolates of Enterococcus faecium resistant to both of these antibiotics. We describe three isolates of vancomycin-resistant E. faecium which demonstrate unique phenotypic characteristics. The isolates exhibited high-level resistance to both vancomycin and teicoplanin, consistent with the VanA phenotype. However, resistance in these isolates could not be induced or cured, and mating experiments failed to detect a transfer of resistance. The combination of vancomycin and penicillin did not significantly change the MIC of penicillin for any of the three isolates. Immunoblotting with polyclonal anti-VanB antibody showed no reaction with the cellular proteins of these strains. Probing with a vanA oligonucleotide revealed hybridization with chromosomal but not plasmid DNA. The mechanism of constitutive resistance of those strains remains unclear. A second mutational change, perhaps involving PBP 5, may explain the presence of resistance to synergistic combination penicillin-vancomycin therapy. In vitro evaluation of penicillin-vancomycin should be carried out in all clinical cases where this therapeutic regimen is being considered. Images PMID:8328775

  6. Porin channels in Escherichia coli: studies with beta-lactams in intact cells.

    PubMed Central

    Nikaido, H; Rosenberg, E Y; Foulds, J

    1983-01-01

    Wild-type Escherichia coli K-12 produces two porins, OmpF (protein 1a) and OmpC (protein 1b). In mutants deficient in both of these "normal" porins, secondary mutants that produce a "new" porin, protein PhoE (protein E), are selected for. We determined the properties of the channels produced by each of these porins by measuring the rates of diffusion of various cephalosporins through the outer membrane in strains producing only one porin species. We found that all porin channels retarded the diffusion of more hydrophobic cephalosporins and that with monoanionic cephalosporins a 10-fold increase in the octanol-water partition coefficient of the solute produced a 5- to 6-fold decrease in the rate of penetration. Electrical charges of the solutes had different effects on different channels. Thus, with the normal porins (i.e., OmpF and OmpC proteins) additional negative charge drastically reduced the penetration rate through the channels, whereas additional positive charge significantly accelerated the penetration. In contrast, diffusion through the PhoE channel was unaffected by the presence of an additional negative charge. We hypothesize that the relative exclusion of hydrophobic and negatively charged solutes by normal porin channels is of ecological advantage to E. coli, which must exclude hydrophobic and anionic bile salts in its natural habitat. The properties of the PhoE porin are also consistent with the recent finding (M. Argast and W. Boos, J. Bacteriol. 143:142-150, 1980; J. Tommassen and B. Lugtenberg, J. Bacteriol. 143:151-157, 1980) that its biosynthesis is derepressed by phosphate starvation; the channel may thus act as an emergency pore primarily for the uptake of phosphate and phosphorylated compounds. Images PMID:6294048

  7. Immunohistochemical mapping of pro-opiomelanocortin- and pro-dynorphin-derived peptides in the alpaca (Lama pacos) diencephalon.

    PubMed

    Manso, B; Sánchez, M L; Medina, L E; Aguilar, L A; Díaz-Cabiale, Z; Narváez, J A; Coveñas, R

    2014-09-01

    Using an indirect immunoperoxidase technique, we studied the distribution of cell bodies and fibres containing non-opioid peptides (adrenocorticotropin hormone (ACTH), alpha-melanocyte-stimulating hormone) and opioid peptides (beta-endorphin (1-27), alpha-neo-endorphin, leucine-enkephalin) in the alpaca diencephalon. No immunoreactive cell bodies containing ACTH were found. Perikarya containing the other four peptides were observed exclusively in the hypothalamus and their distribution was restricted. Perikarya containing alpha-melanocyte-stimulating hormone or alpha-neo-endorphin showed a more widespread distribution than those containing leucine-enkephalin or beta-endorphin (1-27). Cell bodies containing pro-opiomelanocortin-derived peptides were observed in the arcuate nucleus, anterior and lateral hypothalamic areas and in the ventromedial and supraoptic hypothalamic nuclei, whereas perikarya containing alpha-neo-endorphin (a pro-dynorphin-derived peptide) were found in the arcuate nucleus, dorsal and lateral hypothalamic areas, and in the paraventricular, ventromedial and supraoptic hypothalamic nuclei. Immunoreactive cell bodies containing leucine-enkephalin were found in the lateral hypothalamic area and in the paraventricular hypothalamic nucleus. Immunoreactive fibres expressing pro-opiomelanocortin-derived peptides were more numerous than those expressing pro-dynorphin-derived peptides. A close anatomical relationship was observed: in all the diencephalic nuclei in which beta-endorphin (1-27)-immunoreactive fibres were found, fibres containing alpha-melanocyte-stimulating hormone or alpha-neo-endorphin were also observed. Fibres containing beta-endorphin (1-27), alpha-melanocyte-stimulating hormone or alpha-neo-endorphin were widely distributed throughout the diencephalon, but fibres containing ACTH or leucine-enkephalin showed a moderate distribution. The distribution of the five peptides studied here is also compared with that reported previously in

  8. Mathematical model of cAMP-dependent signaling pathway in constitutive and UV-induced melanogenesis

    NASA Astrophysics Data System (ADS)

    Stolnitz, Mikhail M.; Peshkova, Anna Y.

    2002-07-01

    Cascade of reactions of cAMP-dependent signaling pathway in melanocytes is investigated by mathematical modeling. Model takes into account (alpha) -melanocyte stimulating hormone binding to melanocortin-1 receptor, adenylate cyclase activation by G-protein, increase of the intracellular cAMP concentration, PKA activation by cAMP, CREB phosphorylation by PKA, microphthalmia gene expression, microphthalmia binding to tyrosinase gene promoter, increase of tyrosinase synthesis. Positive and negative feedback loops of this system are analyzed.

  9. [N,P]-pyrrole PdCl2 complexes catalyzed the formation of dibenzo-α-pyrone and lactam analogues.

    PubMed

    Suárez-Meneses, J V; Oukhrib, A; Gouygou, M; Urrutigoïty, M; Daran, J-C; Cordero-Vargas, A; Ortega-Alfaro, M C; López-Cortés, J G

    2016-06-21

    We herein report the synthesis and catalytic application of a new family of [N,P] ligands based on the pyrrole ring with alpha-phosphine or phosphole units. Their palladium complexes (3a-d) were obtained in very good yields and their catalytic properties were evaluated in the direct intramolecular arylation to obtain both benzopyranones and phenanthridinones. The air stable complex 3a exhibited the best catalytic performance of this series of complexes, using 1 mol% of catalyst in combination with microwaves to promote this reaction. PMID:27221355

  10. Differential fate of erythromycin and beta-lactam resistance genes from swine lagoon waste under different aquatic conditions.

    PubMed

    Knapp, Charles W; Zhang, Wen; Sturm, Belinda S M; Graham, David W

    2010-05-01

    The attenuation and fate of erythromycin-resistance-methylase (erm) and extended-spectrum beta-lactamse (bla) genes were quantified over time in aquatic systems by adding 20-L swine waste to 11,300-L outdoor mesocosms that simulated receiving water conditions below intensive agricultural operations. The units were prepared with two different light-exposure scenarios and included artificial substrates to assess gene movement into biofilms. Of eleven genes tested, only erm(B), erm(F), bla(SHV) and bla(TEM) were found in sufficient quantity for monitoring. The genes disappeared rapidly from the water column and first-order water-column disappearance coefficients were calculated. However, detected gene levels became elevated in the biofilms within 2 days, but then disappeared over time. Differences were observed between sunlight and dark treatments and among individual genes, suggesting that ecological and gene-specific factors play roles in the fate of these genes after release into the environment. Ultimately, this information will aid in generating better predictive models for gene fate. PMID:20053492

  11. Antimicrobial susceptibility and oxymino-β-lactam resistance mechanisms in Salmonella enterica and Escherichia coli isolates from different animal sources.

    PubMed

    Clemente, Lurdes; Manageiro, Vera; Jones-Dias, Daniela; Correia, Ivone; Themudo, Patricia; Albuquerque, Teresa; Geraldes, Margarida; Matos, Filipa; Almendra, Cláudia; Ferreira, Eugénia; Caniça, Manuela

    2015-09-01

    The impact of extended-spectrum β-lactamases (ESBLs) and plasmid-mediated AmpC β-lactamases (PMAβs) of animal origin has been a public health concern. In this study, 562 Salmonella enterica and 598 Escherichia coli isolates recovered from different animal species and food products were tested for antimicrobial resistance. Detection of ESBL-, PMAβ-, plasmid-mediated quinolone resistance (PMQR)-encoding genes and integrons was performed in isolates showing non-wild-type phenotypes. Susceptibility profiles of Salmonella spp. isolates differed according to serotype and origin of the isolates. The occurrence of cefotaxime non-wild-type isolates was higher in pets than in other groups. In nine Salmonella isolates, blaCTX-M (n = 4), blaSHV-12 (n = 1), blaTEM-1 (n = 2) and blaCMY-2 (n = 2) were identified. No PMQR-encoding genes were found. In 47 E. coli isolates, blaCTX-M (n = 15), blaSHV-12 (n = 2), blaCMY-2 (n = 6), blaTEM-type (n = 28) and PMQR-encoding genes qnrB (n = 2), qnrS (n = 1) and aac(6')-Ib-cr (n = 6) were detected. To the best of our knowledge, this study is the first to describe the presence of blaCMY-2 (n = 2) and blaSHV-12 (n = 1) genes among S. enterica from broilers in Portugal. This study highlights the fact that animals may act as important reservoirs of isolates carrying ESBL-, PMAβ- and PMQR-encoding genes that might be transferred to humans through direct contact or via the food chain. PMID:26054292

  12. 78 FR 22887 - Guidance for Industry on Non-Penicillin Beta-Lactam Drugs: A Current Good Manufacturing Practices...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-17

    ... manufactured, and should have an independent air handling system. A draft version of this guidance was... practices regulation (21 CFR 10.115). The guidance represents the Agency's current thinking on...

  13. Homologous Recombination within Large Chromosomal Regions Facilitates Acquisition of β-Lactam and Vancomycin Resistance in Enterococcus faecium

    PubMed Central

    Lebreton, Francois; McLaughlin, Robert E.; Whiteaker, James D.; Gilmore, Michael S.; Rice, Louis B.

    2016-01-01

    The transfer of DNA between Enterococcus faecium strains has been characterized both by the movement of well-defined genetic elements and by the large-scale transfer of genomic DNA fragments. In this work, we report on the whole-genome analysis of transconjugants resulting from mating events between the vancomycin-resistant E. faecium C68 strain and the vancomycin-susceptible D344RRF strain to discern the mechanism by which the transferred regions enter the recipient chromosome. Vancomycin-resistant transconjugants from five independent matings were analyzed by whole-genome sequencing. In all cases but one, the penicillin binding protein 5 (pbp5) gene and the Tn5382 vancomycin resistance transposon were transferred together and replaced the corresponding pbp5 region of D344RRF. In one instance, Tn5382 inserted independently downstream of the D344RRF pbp5 gene. Single nucleotide variant (SNV) analysis suggested that entry of donor DNA into the recipient chromosome occurred by recombination across regions of homology between donor and recipient chromosomes, rather than through insertion sequence-mediated transposition. The transfer of genomic DNA was also associated with the transfer of C68 plasmid pLRM23 and another putative plasmid. Our data are consistent with the initiation of transfer by cointegration of a transferable plasmid with the donor chromosome, with subsequent circularization of the plasmid-chromosome cointegrant in the donor prior to transfer. Entry into the recipient chromosome most commonly occurred across regions of homology between donor and recipient chromosomes. PMID:27431230

  14. Cephamycins, a new family of beta-lactam antibiotics. I. Production by actinomycetes, including Streptomyces lactamdurans sp. n.

    PubMed

    Stapley, E O; Jackson, M; Hernandez, S; Zimmerman, S B; Currie, S A; Mochales, S; Mata, J M; Woodruff, H B; Hendlin, D

    1972-09-01

    A number of actinomycetes isolated from soil were found to produce one or more members of a new family of antibiotics, the cephamycins, which are structurally related to cephalosporin C. The cephamycins were produced in submerged fermentation in a wide variety of media by one or more of eight different species of Streptomyces, including a newly described species, S. lactamdurans. These antibiotics exhibit antibacterial activity against a broad spectrum of bacteria which includes many that are resistant to the cephalosporins and penicillins. PMID:4790552

  15. Molecular Basis of Filtering Carbapenems by Porins from β-Lactam-resistant Clinical Strains of Escherichia coli.

    PubMed

    Bajaj, Harsha; Scorciapino, Mariano A; Moynié, Lucile; Page, Malcolm G P; Naismith, James H; Ceccarelli, Matteo; Winterhalter, Mathias

    2016-02-01

    Integral membrane proteins known as porins are the major pathway by which hydrophilic antibiotics cross the outer membrane of Gram-negative bacteria. Single point mutations in porins can decrease the permeability of an antibiotic, either by reduction of channel size or modification of electrostatics in the channel, and thereby confer clinical resistance. Here, we investigate four mutant OmpC proteins from four different clinical isolates of Escherichia coli obtained sequentially from a single patient during a course of antimicrobial chemotherapy. OmpC porin from the first isolate (OmpC20) undergoes three consecutive and additive substitutions giving rise to OmpC26, OmpC28, and finally OmpC33. The permeability of two zwitterionic carbapenems, imipenem and meropenem, measured using liposome permeation assays and single channel electrophysiology differs significantly between OmpC20 and OmpC33. Molecular dynamic simulations show that the antibiotics must pass through the constriction zone of porins with a specific orientation, where the antibiotic dipole is aligned along the electric field inside the porin. We identify that changes in the vector of the electric field in the mutated porin, OmpC33, create an additional barrier by "trapping" the antibiotic in an unfavorable orientation in the constriction zone that suffers steric hindrance for the reorientation needed for its onward translocation. Identification and understanding the underlying molecular details of such a barrier to translocation will aid in the design of new antibiotics with improved permeation properties in Gram-negative bacteria.

  16. Effects of β-Lactam Antibiotics and Fluoroquinolones on Human Gut Microbiota in Relation to Clostridium difficile Associated Diarrhea

    PubMed Central

    Heinsen, Femke Anouska; Knecht, Carolin; Schilhabel, Anke; Schmitz, Ruth A.; Zimmermann, Alexandra; dos Santos, Vitor Martins; Ferrer, Manuel; Rosenstiel, Philip C.; Schreiber, Stefan; Friedrichs, Anette K.; Ott, Stephan J.

    2014-01-01

    Clostridium difficile infections are an emerging health problem in the modern hospital environment. Severe alterations of the gut microbiome with loss of resistance to colonization against C. difficile are thought to be the major trigger, but there is no clear concept of how C. difficile infection evolves and which microbiological factors are involved. We sequenced 16S rRNA amplicons generated from DNA and RNA/cDNA of fecal samples from three groups of individuals by FLX technology: (i) healthy controls (no antibiotic therapy); (ii) individuals receiving antibiotic therapy (Ampicillin/Sulbactam, cephalosporins, and fluoroquinolones with subsequent development of C. difficile infection or (iii) individuals receiving antibiotic therapy without C. difficile infection. We compared the effects of the three different antibiotic classes on the intestinal microbiome and the effects of alterations of the gut microbiome on C. difficile infection at the DNA (total microbiota) and rRNA (potentially active) levels. A comparison of antibiotic classes showed significant differences at DNA level, but not at RNA level. Among individuals that developed or did not develop a C. difficile infection under antibiotics we found no significant differences. We identified single species that were up- or down regulated in individuals receiving antibiotics who developed the infection compared to non-infected individuals. We found no significant differences in the global composition of the transcriptionally active gut microbiome associated with C. difficile infections. We suggest that up- and down regulation of specific bacterial species may be involved in colonization resistance against C. difficile providing a potential therapeutic approach through specific manipulation of the intestinal microbiome. PMID:24586762

  17. A cell wall damage response mediated by a sensor kinase/response regulator pair enables beta-lactam tolerance

    PubMed Central

    Dörr, Tobias; Alvarez, Laura; Delgado, Fernanda; Davis, Brigid M.; Cava, Felipe; Waldor, Matthew K.

    2016-01-01

    The bacterial cell wall is critical for maintenance of cell shape and survival. Following exposure to antibiotics that target enzymes required for cell wall synthesis, bacteria typically lyse. Although several cell envelope stress response systems have been well described, there is little knowledge of systems that modulate cell wall synthesis in response to cell wall damage, particularly in Gram-negative bacteria. Here we describe WigK/WigR, a histidine kinase/response regulator pair that enables Vibrio cholerae, the cholera pathogen, to survive exposure to antibiotics targeting cell wall synthesis in vitro and during infection. Unlike wild-type V. cholerae, mutants lacking wigR fail to recover following exposure to cell-wall–acting antibiotics, and they exhibit a drastically increased cell diameter in the absence of such antibiotics. Conversely, overexpression of wigR leads to cell slimming. Overexpression of activated WigR also results in increased expression of the full set of cell wall synthesis genes and to elevated cell wall content. WigKR-dependent expression of cell wall synthesis genes is induced by various cell-wall–acting antibiotics as well as by overexpression of an endogenous cell wall hydrolase. Thus, WigKR appears to monitor cell wall integrity and to enhance the capacity for increased cell wall production in response to damage. Taken together, these findings implicate WigKR as a regulator of cell wall synthesis that controls cell wall homeostasis in response to antibiotics and likely during normal growth as well. PMID:26712007

  18. Comparison of in vitro activity of GR 20263, a novel cephalosporin derivative, with activities of other beta-lactam compounds.

    PubMed Central

    Wise, R; Andrews, J M; Bedford, K A

    1980-01-01

    The in vitro activity of GR 20263, a new cephalosporin, was compared primarily with the activities of moxalactam (LY 127935), cefotaxime, cefoxitin, cefuroxime, and cefazolin against 293 clinical isolates of a variety of gram-positive and -negative bacteria. The minimal inhibitory concentrations of GR 20263 for 90% of group isolates were between 0.06 and 0.5 microgram/ml for the Enterobacteriaceae, Haemophilus influenzae, Neisseria gonorrhoeae, and Lancefield group A beta-hemolytic streptococci; 2 micrograms/ml for Pseudomonas aeruginosa; 16 micrograms/ml for Staphylococcus aureus; and in excess of 128 micrograms/ml for Bacteroides fragilis and Lancefield group D streptococci. In comparison with the other agents, GR 20263 was markedly more active against the Enterobacteriaceae than cefuroxime, cefoxitin, and cefazolin, but marginally less active than moxalactam or cofotaxime. Aganist S. aureus, cefazolin was 16-fold and cefotaxime was 4-fold more active than GR 20263 and moxalactam. GR 20263 was eight-fold more active than cefotaxime and moxalactam against P. aeruginosa. PMID:6772097

  19. Crystal structure of tabtoxin resistance protein complexed with acetyl coenzyme A reveals the mechanism for {beta}-lactam acetylation.

    SciTech Connect

    He, H.; Ding, Y.; Bartlam, M.; Sun, F.; Le, Y.; Qin, X.; Tang, H.; Zhang, R.; Joachimiak, A.; Liu, J.; Zhao, N.; Rao, Z.; Biosciences Division; Tsinghua Univ.; Chinese Academy of Science

    2003-01-31

    Tabtoxin resistance protein (TTR) is an enzyme that renders tabtoxin-producing pathogens, such as Pseudomonas syringae, tolerant to their own phytotoxins. Here, we report the crystal structure of TTR complexed with its natural cofactor, acetyl coenzyme A (AcCoA), to 1.55 {angstrom} resolution. The binary complex forms a characteristic 'V' shape for substrate binding and contains the four motifs conserved in the GCN5-related N-acetyltransferase (GNAT) superfamily, which also includes the histone acetyltransferases (HATs). A single-step mechanism is proposed to explain the function of three conserved residues, Glu92, Asp130 and Tyr141, in catalyzing the acetyl group transfer to its substrate. We also report that TTR possesses HAT activity and suggest an evolutionary relationship between TTR and other GNAT members.

  20. Beta-lactam stability in frozen microdilution PASCO MIC panels using strains with known resistance mechanisms as biosensors.

    PubMed

    Valdezate, S; Martínez-Beltrán, J; de Rafael, L; Baquero, F; Cantón, R

    1996-10-01

    The stability of amoxicillin/clavulanate, piperacillin/tazobactam, cefepime, imipenem, and meropenem in PASCO (PASCO System, DIFCO Laboratories, Detroit, MI, USA) frozen microdilution susceptibility panels stored for 16 weeks at -20 degrees C and -70 degrees C was evaluated. The increase in MIC values for the five American-Type Culture Collection (ATCC) quality control strains for susceptibility testing recommended by the National Committee for Clinical Laboratory Standards (NCCLS) and for 13 strains with different well-characterized resistance mechanisms was indicative of bioactivity deterioration. The overall agreement (+/- 1 twofold dilution) at purchase between the MIC values of PASCO frozen microdilution susceptibility panels and the standard agar dilution method was 97.7%. Minimum inhibitory concentration values for the associations of amoxicillin/clavulanate and piperacillin/tazobactam remained unchanged for the study period at -70 degrees C. In contrast, a carbapenem bioactivity decrease was detected only with strains having well-characterized resistance mechanisms from the 12th week onwards. At -20 degrees C, antibiotic deterioration with these latter strains was observed earlier than with ATCC strains: the activity of meropenem and imipenem remained unchanged only for the first 2 weeks, while a loss of activity was detected for amoxicillin/clavulanate and piperacillin/tazobactam at the 7th and 10th week, respectively. Cefepime was highly stable both at -20 degrees C and -70 degrees C. Strains with well-characterized resistance mechanisms should be used in routine quality control studies of antibiotic stability for susceptibility testing panels during the storage period. PMID:8985656