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Refereed/Peer-reviewed Histophilus somni is a prevalent commensal organism of the upper respiratory tract of cattle and a major causative agent of bovine respiratory disease (BRD) and other syndromes including myocarditis and infectious thromboembolic meningoencephalitis. This study investigated the antimicrobial susceptibility and phylogenetic relationships of H. somni isolates obtained from lung, heart, and other tissues at post-mortem as well as nasal mucosa swabs from cases of BRD in Australian feedlots (2004–2019). Broth microdilution Minimal Inhibitory Concentration (MIC) assays were determined for 19 antimicrobials using three different media (CLSI approved Veterinary Fastidious Medium [VFM], Mueller-Hinton fastidious broth medium supplemented with yeast extract [MHF-Y] and Columbia Broth [CB] supplemented with 5% lysed horse blood). For all antimicrobials, MICs obtained using CB medium were identical or within 1 dilution step of the MICs obtained for VFM and MHF-Y media. Therefore, CB may be a suitable medium for H. somni antimicrobial susceptibility testing similar to MHF-Y medium. None of the 70 Australian H. somni isolates exhibited resistance to antimicrobials with CLSI breakpoints including those commonly used in the treatment of BRD in Australia (first-line tetracyclines [chlortetracycline and oxytetracycline], second-line macrolides [tulathromycin], and third-line extended-spectrum cephalosporin [ceftiofur]). Whole-genome sequence analysis of 65 H. somni isolates for genomic single nucleotide polymorphism differences identified four phylogenetic clusters, each containing isolates from different Australian states, feedlots and tissue sources that clustered together. These findings demonstrate limited genetic diversity and the absence of significant antimicrobial resistance among Australian isolates of H. somni isolated from feedlot cattle.

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Desymmetrisation of robenidine (1: N ',2-bis((E)-4-chlorobenzylidene)hydrazine-1-carboximidhydrazide) and the introduction of imine alkyl substituents gave good antibiotic activity. Of note was the increased potency of two analogues against vancomycin-resistant Enterococci (VRE), one of which returned a MIC of 0.5 mu g mL(-1). Five analogues were found to be equipotent or more potent than the lead 1. Introduction of an indole moiety resulted in the most active robenidine analogue against methicillin-resistant S. aureus (MRSA), with a MIC of 1.0 mu g mL(-1). Imine C=NH isosteres (C=O/C=S) were inactive. Monomeric analogues were 16-64 mu g mL(-1) active against MRSA and VRE. An analogue that lacks the terminal hydrazide NH moiety showed modest Gram-negative activity at 64 mu g mL(-1). A 4-tert-butyl analogue was shown to be active against both Gram-positive and -negative strains at 16-64 mu g mL(-1). In general, additional modifications with aromatic moieties was poorly tolerated, except with concomitant introduction of an imine C-alkyl group. The activity of these analogues against MRSA and VRE ranged from 8 mu g mL(-1) to inactive (MIC>128 mu g mL(-1)) with the naphthyl and indole analogues. Gram-negative activity was most promising with two compounds at 16 mu g mL(-1) against E. coli. Against P. aeruginosa, the highest activity observed was with MIC values of 32 mu g mL(-1) with another two analogues. Combined, these findings support the further development of the (E)-2-benzylidenehydrazine-1-carboximidamide scaffold as a promising scaffold for the development of antibiotics against Gram-positive and Gram-negative strains. Refereed/Peer-reviewed

The bacterial species and specific spoilage organisms associated with the Southern Australian King George Whiting (KGW) and Tasmanian Atlantic Salmon (TAS), and the efficacy of a HOCl-containing water-based sanitization product (Electro-Chemically Activated Solution, by ECAS4) in extending the shelf life of KGW and TAS fillets were evaluated. Fillets were washed with an ECAS4 solution containing either 45 ppm or 150 ppm of free chlorine and bacterial species enumerated on selective and non-selective media, followed by identification of pure isolates by 16 S rRNA gene sequencing. The dominant spoilage microbiota in KGW and TAS fillets stored at 4 ± 1 °C were Pseudomonas spp. and Shewanella spp. At either concentration, ECAS4 significantly reduced total bacterial load and specific spoilage organisms on KGW and TAS fillets (approx. 1–2 log colony-forming units) during storage and significantly extended the shelf life of the fillets by 2 and 4 days, respectively. The significant increase in shelf life and quality of fillets was corroborated by raw and cooked sensory evaluation. ECAS4 sanitization could have a significant impact on the overall food industry, translating into health and economic benefits through reduction of food spoilage bacteria and potentially, foodborne pathogens without many of the disadvantages of currently approved biocides Refereed/Peer-reviewed

Refereed/Peer-reviewed Understanding the underlying molecular mechanism of transporter activity is one of the major discussions in structural biology. A transporter can exclusively transport one ion (specific transporter) or multiple ions (general transporter). This study compared categorical and numerical features of general and specific calcium transporters using machine learning and attribute weighting models. To this end, 444 protein features, such as the frequency of dipeptides, organism, and subcellular location, were extracted for general (n = 103) and specific calcium transporters (n = 238). Aliphatic index, subcellular location, organism, Ile-Leu frequency, Glycine frequency, hydrophobic frequency, and specific dipeptides such as Ile-Leu, Phe-Val, and Tyr-Gln were the key features in differentiating general from specific calcium transporters. Calcium transporters in the cell outer membranes were specific, while the inner ones were general; additionally, when the hydrophobic frequency or Aliphatic index is increased, the calcium transporter act as a general transporter. Random Forest with accuracy criterion showed the highest accuracy (88.88% ±5.75%) and high AUC (0.964 ± 0.020), based on 5-fold cross-validation. Decision Tree with accuracy criterion was able to predict the specificity of calcium transporter irrespective of the organism and subcellular location. This study demonstrates the precise classification of transporter function based on sequence-derived physicochemical features.

Stone formation and catheter blockage are major complications of Proteus UTIs. In this study, we investigated the ability of allicin to inhibit P. mirabilis-induced struvite crystallization and catheter blockage using a synthetic bladder model. Struvite crystallization inhibition study was carried out using P. mirabilis lysate as urease enzyme source in synthetic urine (SU). Struvite productions were monitored by phase contrast light microscopy and measurements of pH, Mg 2+ and Ca 2+ precipitation and turbidity. A catheter blockage study was performed in a synthetic bladder model mimicking natural UTI in the presence of allicin at sub-MIC concentrations (MIC = 64 μg/ml). The results of crystallization study showed that allicin inhibited pH rise and consequently turbidity and precipitation of ions in a dose-dependent manner. The results of catheter blockage study showed that allicin at sub-MIC concentrations (2, 4, 8 μg/ml) significantly increased the time for catheter blockage to occur to 61, 74 and 92 h respectively compared to allicin-free control (48 h). In a similar way, the results showed that allicin delayed the increase of SU pH level in bladder model in a dose-dependent manner compared to allicin-free control. The results also showed that following the increase of allicin concentration, Mg 2+ and Ca 2+ deposition in catheters were much lower compared to allicin-free control, further confirmed by direct observation of the catheters’ eyehole and cross sections. We conclude that allicin prevents the formation of Proteus-induced urinary crystals and the blockage of catheters by delaying pH increase and lowering Mg 2+ and Ca 2+ deposition in a dose-dependent manner. Refereed/Peer-reviewed

Robenidine (E)‐N′‐((E)‐1‐(4‐chlorophenyl)ethylidene)‐2‐(1‐(4‐chlorophenyl)ethylidene)hydrazine‐1‐carboximidhydrazide displays methicillin‐resistant Staphyoccoccus aureus (MRSA) and vancomycin‐resistant Enterococci (VRE) MICs of 2 μg mL−1. Herein we describe the structure‐activity relationship development of a novel series of guanidine to 2‐aminopyrimidine isosteres that ameliorate the low levels of mammalian cytotoxicity in the lead compound while retaining good antibiotic activity. Removal of the 2‐NH2 pyrimidine moiety renders these analogues inactive. Introduction of a central 2‐NH2 triazine moiety saw a 10‐fold activity reduction. Phenyl to cyclohexyl isosteres were inactive. The 4‐BrPh and 4‐CH3Ph with MIC values of 2 and 4 μg mL−1, against MRSA and VRE respectively, are promising candidates for future development.

Objectives Currently, there is no consensus protocols regarding the combination of glycerol (GLY), gelatin or foetal bovine serum (FBS) with dimethyl sulphoxide (DMSO) as cryoprotectants for Mycoplasma bovis in bovine milk samples. This study aimed to compare different cryopreservation compounds and storage temperatures for M. bovis. Results There were significant differences in the survival of M. bovis on different media. Differences were also observed between different storage conditions. All additives improved the survival of M. bovis in comparison to control (CON). The combination of GLY and DMSO was shown to be significantly different to CON with 57.1% (95% CI = 21.43–133.34) and 19.1% (95% CI = 11.73–60.27), respectively at week 16, and its use should be encouraged as a cryoprotectant for M. bovis at − 20 and − 80 °C. GEL/DMSO showed the highest survival rate for M. bovis with 57.14% (95% CI = 21.43–133.34) at 4 °C in comparison with CON 14.29% (95% CI = 9.60–50.39). FBS/DMSO showed the highest survival rate for the short-term preservation similarly to other additives. The evaluated cryopreservative compounds would improve survivability of M. bovis in milk for both transport and long-term storage. Hence, it is recommended to use the mentioned methods for routine transportation or storage purposes for suspicious M. bovis milk samples.

Background Mycoplasma mastitis is increasingly posing significant impact on dairy industry. Although the effects of major conventional mastitis pathogens on milk components has been widely addressed in the literature, limited data on the effects of different Mycoplasma and Acholeplasma spp. on milk quality and quantity is available. The aim of this study was to determine the casual relationship of Mycoplasma spp. and A. laidlawii to mastitis and compare them to subclinical mastitis caused by conventional mastitis pathogens from a single dairy herd in South Australia; Mycoplasma spp. and A. laidlawii were detected using PCR applied directly to milk samples. The herd had mastitis problem with high somatic cell count and low response rate to conventional antimicrobial therapy. A total of 288 cow-level milk samples were collected aseptically and used in this study. Results Conventional culture showed a predominance of coagulase-negative staphylococci, followed by coagulase-positive staphylococci, Streptococcus spp., Enterococcus spp., E. coli, and Klebsiella spp. PCR results showed a high prevalence of mycoplasmas (76.7%), including A. laidlawii (10.8%), M. bovis (6.2%), M. bovirhinis (5.6%), M. arginini (2%), and (52.1%) of cows were co-infected with two or more Mycoplasma and Acholeplasma species. Mycoplasma co-infection significantly increased somatic cell counts (SCC) similar to conventional mastitis pathogens and compared to non-infected cows with 389.3, 550.3 and 67.3 respectively; and decreased the milk yield with 29.0, 29.9 and 34.4 l, respectively. Mycoplasma co-infection caused significant increase in protein percentage, and significant decrease in fat percentage and total milk solids, similar to other conventional mastitis pathogens. In contrast, changes in milk composition and yield caused by various individual Mycoplasma species were non-significant. Conclusions Mycoplasma mastitis had on-farm economic consequences similar to common conventional mastitis pathogens. Results of our study indicate that co-infection Mycoplasma mastitis caused similar effect on milk composition to other mastitis pathogens and we hope these findings raise the awareness of the importance of their detection on routine diagnostic panels.

Sam Abraham, Stephen Jagoe, Stanley Pang, Geoffrey W.Coombs, Mark O'Dea, Jonathon Kelly, Manouchehr Khazandi, Kiro R.Petrovski, Darren J.Trott

The emergence of antimicrobial resistance represents a serious human and animal health risk. Good antimicrobial stewardship is essential to prolong the lifespan of existing antibiotics, and new strategies are required to combat infections in man and animals.To determine the in vitro interaction of ionophores (narasin or monensin) with antimicrobial adjuvants (N-acetylcysteine (NAC), Tris-EDTA or disodium EDTA) against bacterial strains representing pathogens associated with canine otitis externa (OE).American Type Culture Collection (ATCC) strains Staphylococcus aureus 29213, Pseudomonas aeruginosa 27853 and P. aeruginosa biofilm producer PAO1, and a clinical isolate of Proteus mirabilis from a case of canine OE were tested.A 2D microdilution checkerboard method was used, allowing calculation of fractional inhibitory concentration index (FICI), dose reduction index (DRI) and plotting of isobolograms.The combination of narasin with either Tris-EDTA or disodium EDTA produced additive effects (FICI = 0.75) against P. aeruginosa ATCC 27853 and P. aeruginosa biofilm producer ATCC PAO1. An additive effect (FICI = 0.53-0.75) was found against S. aureus ATCC 29213 when narasin or monensin were combined with NAC. The highest DRI (32-fold) was found with monensin/NAC where the MIC of monensin was reduced from 4 to 0.125 μg/mL.The combination of narasin with Tris-EDTA or disodium EDTA is a promising strategy to inhibit the intrinsic resistance elements of Gram-negative bacteria. These novel combinations potentially could be useful as a multimodal approach to treat mixed infections in canine OE.L’émergence de résistances aux antimicrobiens représente un risque pour la santé humaine et animale. Une bonne utilisation des antimicrobiens est essentielle pour prolonger la durée de vie des antibiotiques actuels et de nouvelles stratégies sont nécessaires pour combattre les infections chez l'homme et l'animal. HYPOTHÈSES/OBJECTIFS: Déterminer les interactions in vitro des ionophores (narasine ou monensine) et des adjuvants antimicrobiens (NAC N-acetylcystéine, Tris-EDTA ou disodium EDTA) contre les souches bactériennes pathogènes associées aux otites canines externes (OE).Les souches de l’ATCC (American Type Culture Collection) Staphylococcus aureus 29213, Pseudomonas aeruginosa 27853 et P. aeruginosa productrice de biofilm PAO1, et une souche clinique de Proteus mirabilis issue d'un cas d’OE canine ont été testées. MATÉRIELS ET MÉTHODES: Une méthode de microdilution 2D en damier a été utilisée, permettant un calcul de FICI (fractional inhibitory concentration index), de l'index de réduction de dose (DRI) et des marqueurs d'isobologrammes. RÉSULTATS: L'association de narasine, soit avec Tris-EDTA soit avec disodium EDTA, a produit des effets additifs (FICI = 0.75) contre P. aeruginosa ATCC 27853 et P. aeruginosa producteur de biofilm ATCC PAO1. Un effet additif (FICI = 0.53-0.75) a été trouvé contre S. aureus ATCC 29213 lorsque la narasine ou la monensine étaient combinés avec NAC. Le plus haut DRI (32 fois) a été trouvé avec monensine/NAC là ou le MIC était diminué de 4 à 0.125 μg/mL.L'association de la narasine avec le Tris-EDTA ou le disodium EDTA est une stratégie prometteuse pour inhiber les éléments de résistance des bactéries Gram-négatives. Ces nouvelles associations peuvent potentiellement être utiles dans une approche multimodale pour le traitement des infections mixtes des OE canines.INTRODUCCIÓN: la aparición de resistencia a los antimicrobianos representa un grave riesgo para la salud humana y animal. Una razonable administración de antimicrobianos es esencial para prolongar la vida útil de los antibióticos existentes, y se requieren nuevas estrategias para combatir las infecciones en seres hunanos y animales. HIPÓTESIS/OBJETIVOS: determinar la interacción in vitro de los ionóforos (narasina o monensina) con adyuvantes antimicrobianos (N-acetilcisteína (NAC), Tris-EDTA o EDTA disódico) contra cepas bacterianas que representan patógenos asociados con la otitis externa canina (OE). ANIMALES/AISLADOS: cepas de la American Type Culture Collection (ATCC) Staphylococcus aureus 29213, Pseudomonas aeruginosa 27853 y P. aeruginosa productor de biopelículas PAO1, y un aislado clínico de Proteus mirabilis de un caso de OE canina. MÉTODOS Y MATERIALES: se usó un método de tablero de cuadrículas de microdilución 2D, que permite calcular el índice de concentración inhibitoria fraccional (FICI), el índice de reducción de dosis (DRI) y el trazado de isobologramas. RESULTADOS: la combinación de narasina con Tris-EDTA o EDTA disódico produjo efectos aditivos (FICI = 0,75) frente a P. aeruginosa ATCC 27853 y P. aeruginosa productor de biopelículas ATCC PAO1. Se encontró un efecto aditivo (FICI = 0,53-0,75) frente a S. aureus ATCC 29213 cuando se combinaron narasina o monensina con NAC. El DRI más alto (32 veces) se encontró con monensina/NAC donde la MIC se redujo de 4 a 0,125 μg/ml. CONCLUSIONES E IMPORTANCIA CLÍNICA: la combinación de narasina con Tris-EDTA o EDTA disódico es una estrategia prometedora para inhibir los elementos de resistencia intrínseca de las bacterias Gram-negativas. Estas nuevas combinaciones podrían ser útiles como un enfoque multimodal para tratar infecciones mixtas en la OE canina.Das Auftreten von antimikrobieller Resistenz bedeutet ein schwerwiegendes Gesundheitsrisiko für Mensch und Tier. Eine gute antimikrobielle Stewardship ist essentiell, um die Lebensdauer existierender Antibiotika zu verlängern, außerdem sind neue Strategien nötig, um Infektionen bei Mensch und Tier zu bekämpfen.Die Bestimmung der in vitro Interaktion von Ionophoren (Narasin oder Monensin) mit antimikrobiellen Adjuvantien (N-Acetylcystein (NAC), Tris-EDTA oder Disodium EDTA) gegenüber Bakterienstämmen, die Pathogene darstellen, die bei einer Otitis externa (OE) des Hundes gerne auftreten.Es wurden Stämme aus der American Type Culture Collection (ATCC) von Staphylococcus aureus 29213, Pseudomonas aeruginosa 27853 and P. aeruginosa Biofilm Produzent PAO1, sowie ein klinisches Isolat von Proteus mirabilis von einem Hund mit OE getestet.Es wurde eine 2D Mikrodilutionsschachbrettmethode verwendet, welche die Kalkulation des fraktionellen Hemmstoffkonzentrationsindex (FICI), des Dosisreduktionsindex (DRI) und ein Plotting des Isobologramms erlaubte.Die Kombination von Narasin mit entweder Tris-EDTA oder Disodium EDTA zeigte additive Wirkung (FICI = 0,75) gegen P. aeruginosa ATCC 27853 und P. aeruginosa Biofilm Produzent ATCC PAO1. Eine additive Wirkung (FICI = 0,53-0,75) wurde gegen S. aureus ATCC 29213 gefunden, wenn Narasin oder Monensin mit NAC kombiniert worden waren. Die höchste DRI (32-fach) wurde mit Monensin/NAC gefunden, wodurch die MIC von 4 auf 0,125 µg/mL reduziert wurde.Die Kombination von Narasin mit Tris-EDTA oder Disodium EDTA ist eine vielversprechende Strategie, um die Elemente der intrinsischen Resistenz von Gram-negativen Bakterien zu inhibieren. Diese neuen Kombinationen könnten möglicherweise als multimodale Herangehensweise nützlich sein, um Mischinfektionen bei der OE des Hundes zu behandeln.背景: 抗菌薬耐性の出現は、人間と動物の深刻な健康リスクを表している。既存の抗生物質のライフスパンを延ばすためには、優れた抗菌薬管理が不可欠であり、人間や動物の感染症と闘うには新しい戦略が必要である。 仮説/目的: 本研究の目的は、犬外耳炎(OE)に関連する病原体を表す細菌株に対するイオノフォア(ナラシンまたはモネンシン)および抗菌アジュバント(N-アセチルシステイン(NAC)、Tris-EDTAまたはEDTA二ナトリウム)のin vitroにおける相互作用を決定することである。 被験動物/分離株: 黄色ブドウ球菌American Type Culture Collection(ATCC)29213、緑膿菌ATCC 27853および緑膿菌バイオフィルム産生株ATCC PAO1、および犬外耳炎症例からProteus mirabilisの臨床分離株を検査した。 材料と方法: 2D微量希釈チェッカーボード法を使用して、分別阻害濃度指数(FICI)、線量低減指数(DRI)の計算およびアイソボログラム法のプロッティングを可能にした。 結果: ナラシンとTris-EDTAまたはEDTA二ナトリウムの併用は、緑膿菌ATCC 27853および緑膿菌バイオフィルム産生株ATCC PAO1に対して相加効果(FICI = 0.75)をもたらした。ナラシンまたはモネンシンをNACと併用した場合、黄色ブドウ球菌ATCC 29213に対して相加効果(FICI = 0.53-0.75)を認めた。最も高いDRI(32倍)は、MICが4から0.125μg/ mLに減少したモネンシン/ NACで検出された。 結論と臨床的重要性: ナラシンとTris-EDTAまたは二ナトリウムEDTAの併用は、グラム陰性菌固有の耐性要素を抑制する有望な戦略である。これらの新しい併用法は、犬外耳炎の混合感染症治療のマルチモーダルアプローチとして有用である可能性がある。.背景: 抗菌素耐药问题会带来人类和动物严重的健康风险。良好的抗菌素管理对于延长现有抗生素的产品寿命至关重要,需要采取新的策略来对抗人和动物的感染。 假设/目的: 确定离子型抗生素(那拉菌素或莫能菌素)与抗微生物佐剂(N-乙酰半胱氨酸(NAC),Tris-EDTA或EDTA二钠)相互作用后,抗犬外耳炎(OE)病原体的体外效果。 动物/分离株: 测试美国菌种保藏中心(ATCC)的金黄色葡萄球菌29213,铜绿假单胞菌27853和铜绿假单胞菌生物膜形成物PAO1,以及从犬OE病例中分离出的变形杆菌。 方法和材料: 采用2D微稀释棋盘法,计算分级抑制浓度指数(FICI)、剂量降低指数(DRI)并绘制等效线图。 结果: 那拉菌素与Tris-EDTA或EDTA二钠的组合,对铜绿假单胞菌ATCC 27853和铜绿假单胞菌生物膜形成物ATCC PAO1具有协同作用(FICI = 0.75)。 当那拉菌素或莫能菌素与NAC联用时,发现对金黄色葡萄球菌ATCC 29213同样具有协同作用(FICI = 0.53-0.75)。 莫能菌素/ NAC的DRI最高(32倍),MIC从4降至0.125μg/ mL。 结论和临床意义: 对抑制革兰氏阴性细菌的固有耐药因素,那拉菌素联合Tris-EDTA或EDTA二钠是一种有前途的策略。 这些新颖的联合,可作为犬OE混合感染的多模式治疗方案。.O surgimento de resistência antimicrobiana representa um sério risco à saúde humana e animal. Boas práticas de administração de antimicrobianos são essenciais para prolongar a vida útil dos antibióticos existentes, e novas estratégias são necessárias para combater infecções no homem e nos animais. HIPÓTESE/OBJETIVOS: Determinar a interação in vitro de ionóforos (narasina ou monensina) com adjuvantes antimicrobianos (N-acetilcisteína (NAC), Tris-EDTA ou EDTA dissódico) contra cepas bacterianas associados à otite externa canina (OE).As cepas padrão registradas na American Type Culture Collection (ATCC) de Staphylococcus aureus 29213, Pseudomonas aeruginosa 27853, P. aeruginosa PAO1 produtora de biofilme, e um isolado clínico de Proteus mirabilis oriundo de um caso de OE canino foram testados. MÉTODOS E MATERIAIS: Utilizou-se o método de checkerboard em microdiluição 2D, permitindo o cálculo do índice de concentração inibitória fracionária (FICI), índice de redução da dose (DRI) e plotagem de isobologramas.A combinação de narasina com Tris-EDTA ou EDTA dissódico prossuiu efeitos aditivos (FICI = 0,75) contra P. aeruginosa ATCC 27853 e P. aeruginosa ATCC PAO1 produtora de biofilme. Um efeito aditivo (FICI = 0,53-0,75) foi encontrado contra S. aureus ATCC 29213 quando narasina ou monensina foram combinadas com NAC. O maior DRI (32 vezes) foi encontrado com monensina / NAC, onde a CIM foi reduzida de 4 para 0,125 μg / mL. CONCLUSÕES E IMPORTÂNCIA CLÍNICA: A combinação de narasina com Tris-EDTA ou EDTA dissódico é uma estratégia promissora para inibir os elementos intrínsecos de resistência de bactérias Gram-negativas. Essas novas combinações poderiam ser potencialmente úteis como uma abordagem multimodal no tratamento de infecções mistas nas OEs caninas.

An antibiotic adjuvant is a chemical substance used to modify or augment the effectiveness of primary antimicrobial agents against drug-resistant micro-organisms. Its use provides an alternative approach to address the global issue of antimicrobial resistance and enhance antimicrobial stewardship.To determine the antimicrobial activity of a panel of potential antimicrobial adjuvants against common pathogens associated with canine otitis externa (OE).A number of type strains and clinical isolates (n = 110) from canine OE were tested including Staphylococcus pseudintermedius, β-haemolytic Streptococcus spp., Pseudomonas aeruginosa, Proteus mirabilis and Malassezia pachydermatis.Antimicrobial activities of monolaurin, monocaprin, N-acetylcysteine (NAC), polymyxin B nonapeptide, Tris-EDTA, Tris-HCL and disodium EDTA were tested using microdilution methodology according to CLSI guidelines.N-acetylcysteine, Tris-EDTA and disodium EDTA had antimicrobial activity against both type strains and otic pathogens. The other adjuvants tested had limited to no efficacy. NAC had a minimal inhibitory concentration (MIC) range of 2,500-10,000 μg/mL for the various organisms. Pseudomonas aeruginosa isolates were eight times more susceptible to disodium EDTA in the presence of Tris-HCL in comparison to disodium EDTA alone. Malassezia pachydermatis isolates were most susceptible to Tris-EDTA (MICN-acetylcysteine, Tris-EDTA and disodium EDTA have intrinsic antimicrobial activity and represent promising adjuvants that could be used to enhance the efficacy of existing antibiotics against Gram-negative and multidrug-resistant bacterial infections. These agents could be combined with other antimicrobial agents in a multimodal approach for mixed ear infections in dogs.Un adjuvant antibiotique est une substance chimique utilisée pour modifier ou augmenter l'efficacité d'agents antimicrobiens primaires contre les micro-organismes résistants; Leur utilisation fournit une approche alternative pour gérer les résistances antimicrobiennes et augmenter la gestion antimicrobienne. HYPOTHÈSES/OBJECTIFS: Déterminer l'activité antimicrobienne d'un panel d'adjuvants antimicrobien potentiels contre les pathogènes fréquents associés aux otites externes canines (OE).Un nombre de souches types et de prélèvements cliniques (n = 110) d'OE canines ont été testés comprenant Staphylococcus pseudintermedius, β-haemolytic Streptococcus spp., Pseudomonas aeruginosa, Proteus mirabilis et Malassezia pachydermatis. MÉTHODES: Les activités antimicrobiennes de monolaurine, monocaprine, N-acetylcystéine (NAC), polymyxin B nonapeptide, Tris-EDTA, Tris-HCL et disodium EDTA ont été testées par microdilution selon les recommandations du CLSI. RÉSULTATS: Les N-acetylcystéine, Tris-EDTA et disodium EDTA avaient une activité antimicrobienne contre les deux types de souches et les pathogènes auriculaires. Les autres adjuvants testés avaient une efficacité limitée à pas d'efficacité. NAC avait une concentration minimale inhibitrice (MIC) qui variait de 2,500 à 10,000 μg/mL pour les différents organismes. Les souches de Pseudomonas aeruginosa étaient huit fois plus sensibles au disodium EDTA en présence de Tris-HCL en comparaison au disodium EDTA seul. Les souches de Malasseiza pachydermatis étaient plus sensibles au Tris-EDTA (MICN-acetylcystéine, Tris-EDTA et disodium EDTA avaient une activité antimicrobienne intrinsèque et représentaient des adjuvants prometteurs qui pourraient être utilisés pour augmenter l'efficacité des antibiotiques existants contre les infections bactériennes gram négatives et multi-résistantes. Ces agents pourraient être associés avec d'autres agents antimicrobiens dans une approche multimodale des infections de l'oreille du chien.INTRODUCCIÓN: un antibiótico adyuvante es una sustancia química utilizada para modificar o aumentar la efectividad de los agentes antimicrobianos primarios contra microorganismos resistentes a los medicamentos. Su uso proporciona un enfoque alternativo para abordar el problema global de la resistencia a los antimicrobianos y mejorar la administración antimicrobiana. HIPÓTESIS/OBJETIVOS: determinar la actividad antimicrobiana de un panel de potenciales adyuvantes antimicrobianos contra patógenos comunes asociados con la otitis externa canina (OE). ANIMALES/AISLAMIENTOS: se analizaron cepas tipo y aislamientos clínicos (n = 110) de OE canina, que incluían Staphylococcus pseudintermedius, Streptococcus spp. β-hemolítico, Pseudomonas aeruginosa, Proteus mirabilis y Malassezia pachydermatis. MÉTODOS: se analizaron las actividades antimicrobianas de monolaurina, monocaprina, N-acetilcisteína (NAC), polimixina B nonapéptido, Tris-EDTA, Tris-HCL y EDTA disódico utilizando la metodología de microdilución de acuerdo con las pautas CLSI. RESULTADOS: N-acetilcisteína, Tris-EDTA y EDTA disódico tuvieron actividad antimicrobiana contra cepas tipo y patógenos óticos aislados de campo. Los otros adyuvantes probados tuvieron eficacia limitada o no existente. La NAC tenía un rango de concentración inhibitoria mínima (CIM) de 2,500 a 10,000 μg/ml para los diversos organismos. Los aislamientos de Pseudomonas aeruginosa fueron ocho veces más susceptibles al EDTA disódico en presencia de Tris-HCL en comparación con el EDTA disódico solo. Los aislados de Malassezia pachydermatis fueron más susceptibles a Tris-EDTA (MIC90 = 190/60 μg/mL) y EDTA disódico (MIC90 = 120 μg/mL). CONCLUSIONES Y RELEVANCIA CLÍNICA: la N-acetilcisteína, Tris-EDTA y EDTA disódico tienen actividad antimicrobiana intrínseca y representan adyuvantes prometedores que podrían usarse para mejorar la eficacia de los antibióticos existentes contra las infecciones bacterianas Gram-negativas y resistentes a múltiples fármacos. Estos agentes podrían combinarse con otros agentes antimicrobianos en un enfoque multimodal para infecciones de oído mixtas en perros.Ein antibiotisches Adjuvans ist eine chemische Substanz, die eingesetzt wird, um die Wirksamkeit von primär antibiotischen Wirkstoffen gegenüber Medikamenten-resistenten Mikroorganismen zu modifizieren oder zu verbessern. Seine Verwendung bedeutet eine alternative Herangehensweise, um das globale Thema der antimikrobiellen Resistenz anzusprechen und Antimikrobielle Stewardship zu verbessern.Die Bestimmung der antimikrobiellen Aktivität eines Panels von potentiellen antimikrobiellen Adjuvantien gegen übliche Pathogene, die mit einer Otitis externa (OE) des Hundes in Zusammenhang gebracht werden.Eine Anzahl von Erregerstämmen und klinische Isolate (n = 110) von OE von Hunden mit Staphylococcus pseudintermedius, β-haemolytic Streptococcus spp., Pseudomonas aeruginosa, Proteus mirabilis und Malassezia pachydermatis.N-Acetylcystein, Tris-ETDA und Disodium EDTA zeigen eine antimikrobielle Aktivität gegenüber beiden Stämmen und Ohr Pathogenen. Die anderen getesteten Adjuvantien zeigten wenig bis keine Wirksamkeit. NAC hatte eine minimale Hemmkonzentration (MIC) im Bereich von 2,500-10,000 μg/mL für die verschiedenen Organismen. Pseudomonas aeruginosa Isolate waren achtmal empfänglicher auf Disodium EDTA wenn auch Tris-HCl verwendet wurde im Vergleich zu Disodium EDTA alleine. Malassezia pachydermatis Isolate waren am empfindlichsten auf Tris-EDTA (MICN-Acetylcystein, Tris-EDTA und Disodium EDTA haben eine intrinsische antimikrobielle Aktivität und repräsentieren vielversprechende Adjuvantien, die verwendet werden könnten, um die Wirksamkeit von bestehenden Antibiotika gegenüber Gram-negativen und Multidrug-resistenten bakteriellen Infektionen zu verbessern. Diese Wirkstoffe könnten mit anderen antimikrobiellen Wirkstoffen in einer multimodalen Herangehensweise für Mischinfektionen der Ohren von Hunden kombiniert werden.背景: 抗菌アジュバントは、薬剤耐性菌に対する主要な抗菌薬の有効性を修飾または増強するために使用される化学物質である。抗菌アジュバントの使用は、世界的な抗生剤耐性問題に対処し、抗菌管理を強化するための代替アプローチとして提供される。 仮説/目的: 本研究の目的は、犬の外耳炎(OE)に関連する一般的な病原体に対する潜在的な抗菌アジュバントパネルの抗菌活性を決定することである。 被験動物/分離株: 犬のOE由来の多くの基準株および臨床分離株(n = 110)(Staphylococcus pseudintermedius、β-haemolytic Streptococcus spp.、Pseudomonas aeruginosa、Proteus mirabilisおよびMalassezia pachydermatis)を試験した。 方法: monolaurin、monocaprin、N-acetylcysteine (NAC)、polymyxin B nonapeptide、Tris-EDTA、Tris-HCLおよびdisodium EDTAの抗菌活性を、CLSIガイドラインに従って微量希釈法を用いて試験した。 結果: N-acetylcysteine、Tris-EDTAおよびdisodium EDTAは、基準株および耳病原体の両方に対して抗菌活性を有していた。試験した他のアジュバントの有効性は限定的であった。 NACは、様々な微生物に対して、2,500〜10,000μg/ mLの最小阻害濃度(MIC)範囲を有していた。 Pseudomonas aeruginosa分離株は、disodium EDTA単独と比較して、Tris-HCl存在下でdisodium EDTAに8倍の感受性を示した。 Malassezia pachydermatis分離株は、Tris-EDTA(MIC背景: 抗生素佐剂是一种化学物质,用于改变或增强主要抗菌剂对抗耐药微生物的有效性。它的使用为解决全球抗菌素耐药性问题,以及加强抗菌药物管理提供了一种备选方法。 假设/目的: 确定一组潜在抗菌佐剂对犬外耳炎(OE)常见病原体的抗菌活性。 动物/分离物: 对多种来自犬OE的不同类型的正常和致病菌株进行测试(n = 110),包括假中间型葡萄球菌、β-溶血性链球菌、铜绿假单胞菌、奇异变形杆菌和厚皮马拉色菌。 方法: 根据CLSI指南,采用微量稀释方法检测月桂酸甘油酯、单癸酸甘油脂、N-乙酰半胱氨酸(NAC),多粘菌素B非肽、Tris-EDTA、Tris-HCL和EDTA二钠的抗菌活性。 结果: N-乙酰半胱氨酸,Tris-EDTA和EDTA二钠对两种类型菌株和耳病原体都具有抗菌活性;测试的其他佐剂功效有限。对于各种微生物,NAC最小抑制浓度(MIC)范围是2,500-10,000μg/ mL。与单独的EDTA二钠相比,在Tris-HCL存在下,铜绿假单胞菌株对EDTA二钠的敏感性高8倍。厚皮马拉色菌株对Tris-EDTA(MIC90 = 190 /60μg/ mL)和EDTA二钠(MIC90 =120μg/ mL)最敏感。 结论和临床相关性: N-乙酰半胱氨酸、Tris-EDTA和EDTA二钠具有内在抗菌活性,是一种有前途的佐剂,可用于增强现有抗生素对抗革兰氏阴性和多药耐药性细菌感染的功效。这些药剂可以与其他抗微生物剂组合,以多种模式用于治疗犬耳的混合感染。.Um adjuvante de antibiótico é uma substância química utilizada para modificar ou aumentar a eficácia de agentes antimicrobianos primários contra microrganismos resistentes. O seu uso propicia uma alternativa para abordar o problema global da resistência a antimicrobianos e enfatizar as diretrizes para uso racional destes fármacos (antimicrobial stewardship). HIPÓTESE/OBJETIVOS: Determinar a atividade antimicrobiana de um painel de potenciais adjuvantes de antimicrobianos contra patógenos comumente associados à otite externa canina (OE). Animais/Isolados Testou-se cepas padrão e isolados clínicos (n = 110) de OE canina, incluindo Staphylococcus pseudintermedius, Streptococcus spp. β-haemolítico, Pseudomonas aeruginosa, Proteus mirabilis e Malassezia pachydermatis. MÉTODOS: A atividade antimicrobiana de monolaurina, monocaprina, N-acetilcisteína (NAC), nanopeptídeo polimixina B, Tris-eDTA, Tris-HCL e EDTA dissódico foi testada utilizando o método de microdiluição de acordo com as diretrizes do CLSI.N-acetilcisteína, Tris-EDTA e EDTA dissódico apresentaram atividade antimicrobiana tanto contra as cepas padrão quanto contra os patógenos óticos. Os outros adjuvantes apresentaram pouca ou nenhuma eficácia. A NAC apresentou um intervalo de concentração inibitória mínima (MIC) de 2.500-10.000 μg/mL para os diversos microrganismos. Os isolados de Pseudomonas aeruginosa foram oito vezes mais suscetíveis ao EDTA dissódico na presença do Tris-HCL em comparação ao EDTA dissódico isoladamente. Os isolados de Malassezia pachydermatis foram mais suscetíveis ao Tris-EDTA (MIC

Robenidine, 1 (2,2′-bis[(4-chlorophenyl)methylene]carbonimidic dihydrazide), was active against MRSA and VRE with MIC’s of 8.1 and 4.7 μM, respectively. SAR revealed tolerance for 4-Cl isosteres with 4-F (8), 3-F (9), 3-CH3 (22), and 4-C(CH3)3 (27) (23.7–71 μM) and with 3-Cl (3), 4-CH3 (21), and 4-CH(CH3)2 (26) (8.1–13.0 μM). Imine carbon alkylation identified a methyl/ethyl binding pocket that also accommodated a CH2OH moiety (75; 2,2′-bis[1-(4-chlorophenyl)-2-hydroxyethylidene]carbonimidic dihydrazide). Analogues 1, 27 (2,2′-bis{[4-(1,1-dimethylethyl)phenyl]methylene}carbonimidic dihydrazide), and 69 (2,2′-bis[1-(4-chlorophenyl)ethylidene]carbonimidic dihydrazide hydrochloride) were active against 24 clinical MRSA and MSSA isolates. No dose-limiting cytotoxicity at ≥2× MIC or hemolysis at ≥8× MIC was observed. Polymyxin B addition engendered Escherichia coli and Pseudomonas aeruginosa Gram-negative activity MIC’s of 4.2–21.6 μM. 1 and 75 displayed excellent microsomal stability, intrinsic clearance, and hepatic extraction ratios with T1/2 > 247 min, CLint < 7 μL/min/mg protein, and EH < 0.22 in both human and mouse liposomes for 1 and in human liposomes for 75. Refereed/Peer-reviewed

A frozen section method utilising chicken intestinal tissue was developed to study the Eimeria tenella attachment ex vivo. In order to examine Eimeria-epithelial cell attachment, 10(5) E. tenella sporozoites were incubated with each caecal frozen section (6, 10 and 14 microm) for 1h in 5% CO2 incubator at 41 degrees C. E. tenella sporozoites attached successfully to enterocytes in 14 microm thick of caecal sections. Sporozoite attachment to caecal sections was shown to be dependent on the number of parasites added. To evaluate the method, E. tenella sporozoites were incubated to its preferred (caecum) and non-preferred (duodenum and jejunum) intestinal sites. The number of sporozoites attached to the caecal enterocytes was significantly greater (P

Antimicrobial resistance and antimicrobial stewardship are of ever-increasing importance in veterinary medicine. Re-purposing of old drugs that are not used in human medicine is one approach that addresses the emergence of multidrug resistance in canine skin and ear infections, and can reduce the use of critically important human antibiotic classes.To determine the antimicrobial activity of narasin, a polyether ionophore conventionally used as a rumen modifier and anticoccidial agent in production animals, against common clinical isolates of canine otitis externa (OE).Clinical isolates (n = 110) from canine OE were tested, including 17 meticillin-susceptible Staphylococcus pseudintermedius (MSSP), 13 multidrug-resistant Staphylococcus pseudintermedius (MDRSP), and 20 each of β-haemolytic Streptococcus spp., Pseudomonas aeruginosa, Proteus mirabilis and Malassezia pachydermatis.Bacterial and yeast isolates were subcultured, suspended in broth and inoculated into 96-well plates. Organisms were tested against concentrations of narasin ranging from 0.03 to 128 μg/mL. Minimal inhibitory concentrations (MICs) were determined after overnight incubation.Narasin MICs for staphylococcal and streptococcal isolates ranged from 0.06 to 0.25 μg/mL; MICNarasin was effective against Gram-positive bacteria and had antifungal activity at higher concentrations against M. pachydermatis. However, the lack of Gram-negative activity would prevent its use as a sole antimicrobial agent in cases of canine OE.

Developing a reliable mastitis challenge infection model is required to test new intramammary antimicrobial preparations, other novel bovine mastitis treatments, and study mastitis pathogenesis. Three treatment groups of Holstein Friesian cows in active lactation were administered two doses (104 and 106 cfu/quarter) on a single occasion with one of the three Streptococcus uberis strains (BFR6019, MFF1283 and SA002) suspended in 5 ml of sterile PBS, administered via intramammary inoculation immediately after milking. All quarters that were challenged with S. uberis strains MLF1283 and BFR6019 showed clinical signs of mastitis on day 1 and 2 after the challenge. Strain SA002 had a lower rate of inducing clinical mastitis which was detected later than day 3 after the challenge. We successfully developed a rapid and reliable model for inducing experimental S. uberis mastitis with 100% success rate in cows in active lactation. On the basis of the correlation results between strains, RAPD fingerprinting results, clinical findings, and a 100% success rate of mastitis induction for low and high doses S. uberis strains MLF1283 and BFR6019, strain virulence seems to be a more important effect than challenge dose in induction of clinical mastitis following experimental challenge.

Meticillin-resistant Staphylococcus pseudintermedius (MRSP) has recently emerged as a worldwide cause of canine pyoderma. In this study, we characterized 22 S. pseudintermedius isolates cultured from 19 dogs with pyoderma that attended a veterinary dermatology referral clinic in Australia in 2011 and 2012. Twelve isolates were identified as MRSP by mecA real-time PCR and phenotypic resistance to oxacillin. In addition to β-lactam resistance, MRSP isolates were resistant to erythromycin (91.6 %), gentamicin (83.3 %), ciprofloxacin (83.3 %), chloramphenicol (75 %), clindamycin (66 %), oxytetracycline (66 %) and tetracycline (50 %), as shown by disc-diffusion susceptibility testing. Meticillin-susceptible S. pseudintermedius isolates only showed resistance to penicillin/ampicillin (90 %) and tetracycline (10 %). PFGE using the SmaI restriction enzyme was unable to type nine of the 12 MRSP isolates. However the nine isolates provided the same PFGE pulsotype using the Cfr91 restriction enzyme. Application of the mec-associated direct repeat unit (dru) typing method identified the nine SmaI PFGE-untypable isolates as dt11cb, a dru type that has only previously been associated with MRSP sequence type (ST)45 isolates that possess a unique SCCmec element. The dt11cb isolates shared a similar multidrug-resistant antibiogram phenotype profile, whereas the other MRSP isolates, dt11a, dt11af (dt11a-associated) and dt10h, were resistant to fewer antibiotic classes and had distinct PFGE profiles. This is the first report of MRSP causing pyoderma in dogs from Australia. The rapid intercontinental emergence and spread of multidrug-resistant MRSP strains confirms the urgent need for new treatment modalities for recurrent canine pyoderma in veterinary practice.

Periodontal disease is a common disease of dogs and cats often requiring antimicrobial treatment as an adjunct to mechanical debridement. However, correct compliance with oral antimicrobial therapy in companion animals is often difficult. Cefovecin is a recently introduced veterinary cephalosporin that has demonstrated prolonged concentrations in extracellular fluid, allowing for dosing intervals of up to 14 days. Subgingival samples were collected from the oral cavity of 29 dogs and eight cats exhibiting grade 2 or grade 3 periodontal disease. Samples were cultivated on Wilkin Chalgrens agar and incubated in an anaerobic chamber for seven days. Selected anaerobic bacteria were isolated and identified to species level using 16S rRNA gene sequence analysis. Minimum inhibitory concentrations were determined for cefovecin and six additional antimicrobials using the agar dilution methodology recommended by the Clinical and Laboratory Standards Institute. The 65 clinical isolates were identified as Porphyromonas gulae (n = 45), Porphyromonas crevioricanis (n = 12), Porphyromonas macacae (n = 1), Porphyromonas cangingivalis (n = 1) Fusobacterium nucleatum (n = 2), Fusobacterium russii (n = 1) and Solobacterium moorei (n = 3). This is the first report of S. moorei being isolated from companion animals with periodontal disease. All isolates were highly susceptible to cefovecin, with a MIC90 of ≤0.125 μg/ml. Conversely, different resistance rates to ampicillin, amoxicillin and erythromycin between isolates were detected. Cefovecin is thus shown to be effective in vitro against anaerobic bacteria isolated from dogs and cats with periodontal disease.