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One of the major problems to be addressed in aquaculture is the prominence of antimicrobial resistance (AMR). The occurrence of bacterial infections in cultured fishes promotes the continuous use of antibiotics in aquaculture, which results in the selection of proliferated antibiotic‐resistant bacteria and increases the possibility of transfer to the whole environment through horizontal gene transfer. Hence, the accurate cultivation‐dependent and cultivation‐independent detection methods are very much crucial for the immediate and proper management of this menace. Antimicrobial resistance determinants carrying mobile genetic transfer elements such as transposons, plasmids, integrons and gene cassettes need to be specifically analysed through molecular detection techniques. The susceptibility of microbes to antibiotics should be tested at regular intervals along with various biochemical assays and conjugation studies so as to determine the extent of spread of AMR. Advanced omic‐based and bioinformatic tools can also be incorporated for understanding of genetic diversity. The present review focuses on different detection methods to unearth the complexity of AMR in aquaculture. This monitoring helps the authorities to curb the use of antibiotics, commencement of appropriate management measures and adequate substitute strategies in aquaculture. The long battle of AMR could be overcome by the sincere implementation of One Health approach. Significance and Impact of the Study: The use of antibiotics and increased antimicrobial resistance (AMR) are of major concerns in aquaculture industry. This could result in global health risks through direct consumption of cultured fishes and dissemination of AMR to natural environment through horizontal gene transfer. Hence, timely detection of the antimicrobial‐resistant pathogens and continuous monitoring programmes are inevitable. Advanced microbiological, molecular biological and omic‐based tools can unravel the menace to a great extent. This will help the authorities to curb the use of antibiotics and implement appropriate management measures to overcome the threat. [ABSTRACT FROM AUTHOR]

Serogroup 6 remains common in the pneumococcal-conjugated vaccine era in Bulgaria; therefore, we investigated its clonal and serotype dynamics. The antibiotic susceptibilities were assessed by broth microdilution. Strains identified as serogroup 6 with latex agglutination method were subjected to serotype-specific PCRs. Erythromycin-resistant strains were analyzed by PCR for presence of ermB and mefE genes. MLST was performed to define clonal composition of the sequence types (STs). Serogroup 6 was represented by 40 (13.3%) from 301 invasive and non-invasive Streptococcus pneumoniae isolates. Molecular serotyping revealed new emerging serotype 6C (6.6%), not detected in pre-vaccine era. Among unvaccinated patients, mostly we observed serotypes 6А (57.1%) and 6В (28.6%). Serotype 6C was distinctive for vaccinated children (64%), followed by 6A (24%). Penicillin and ceftriaxone non-susceptible serogroup 6 strains were 65% and 5%, respectively; erythromycin- and clindamycin-resistant were 70.0% and 52.5%, respectively. Multidrug-resistant strains were 57.5%. Prevalent genetic determinant for macrolide resistance was ermB gene (75%). MLST revealed 17 STs into 5 clonal complexes and 7 singletons. Predominant genetic lineage was CC386, represented by MDR-6C non-invasive strains. Serotype 6B, principally responsible for invasive diseases in the pre-vaccine era, retreated this position to serotype 6A. [ABSTRACT FROM AUTHOR]

Objective: The current investigation was designed to point out the prevalence of multidrug-resistant Streptococcus spp. causing acute clinical mastitis and their pattern of antibiotic resistance in dairy cows. Materials and methods: Milk was sampled from 128 dairy cows with 191 infected quarters during the period from August 2017 to December 2018. Bacterial species were isolated from the milk samples and identified based on colony morphology and biochemical tests. Multiplex PCR was done for confirmatory detection of the Streptococcus spp. isolates. Results: The chief isolation percentages, from the sampled milk, were Escherichia coli (26%), then Staphylococcus aureus (23%), and Streptococcus dysagalactiae (23%), then Streptococcus agalactiae (20.1%), and finally coagulase-negative Staphylococci (7.7%). In confirmed PCR streptococci isolates, the antibiotic resistance genes have been detected, including macrolides antibiotic resistance genes (ermB and mefA genes), lincosamides antibiotic resistance genes (linB gene), and tetracycline resistance genes (tetM and tetO genes). Age, parity number, cleaning of bedding materials, cleaning of milking facilities, and utensils and udder cleaning practice were significant risk factors for multidrug-resistant streptococcal mastitis in dairy cows. Conclusion: The results of this study explored the phenotypic and genotypic traits of Streptococcus spp. which constitute a usual cause of acute clinical mastitis in dairy cows. The ermB, mefA, tetM, and tetO antibiotic-resistant genes were identified in streptococci isolates from dairy cows' milk with acute clinical mastitis, indicating a public health hazard. Thus, veterinary clinical breakpoints are needed to improve surveillance data, improve the hygiene regimen on the farms, and promote the wise use of antimicrobials. [ABSTRACT FROM AUTHOR]

Background: The surveillance of emm types and macrolide susceptibility of group A streptococcus (GAS) in various areas and time periods enhances the understanding of the epidemiology of GAS infections and may guide treatment strategies and the formulation of type-specific vaccines. Greece has emerged as a country with high macrolide use. However, studies suggest a gradual reduction in macrolide consumption after 2007. Methods: During a 7-year period (2011–2017), 604 GAS isolates were recovered from consecutive children presenting with pharyngeal or nonpharyngeal infections in Central Greece; 517 viable isolates underwent molecular analysis, including emm typing. Results: Isolates belonged to 20 different emm types (in decreasing order of prevalence: 1, 89, 4, 12, 28, 3, 75 and 6, accounting for 88.2% of total isolates). The emm types comprised 10 emm clusters (five most common clusters: E4, A-C3, E1, A-C4 and A-C5). The emm89 isolates were acapsular ('new clade'). Overall macrolide resistance rate was 15.4%, and cMLSB emerged as the predominant resistance phenotype (56.4%). The lowest annual resistance rates occurred in 2014 (13.1%), 2016 (5.5%) and 2017(8.0%) (P for trend = 0.002). Consumption of macrolide/lincosamide/streptogramin B declined by 22.6% during 2011–2017. Macrolide resistance and emm28 and emm77 types were associated (both P<0.001). The most frequently identified genetic lineages of macrolide-resistant GAS included emm28/ST52, emm77/ST63, emm12/ST36, emm89/ST101 and emm4/ST39. We estimated that 98.8% of the isolates belonged to emm types incorporated into a novel 30-valent M protein vaccine. Conclusions: In Central Greece during 2011–2017, the acapsular emm89 isolates comprised the second most prevalent type. Susceptibility testing and molecular analyses revealed decreasing GAS macrolide resistance rates, which may be attributed to the reduction in the consumption of macrolides and/or the reduced circulation of macrolide-resistant clones in recent years. Such data may provide valuable baseline information in targeting therapeutic intervention and the formulation of type-specific GAS vaccines. [ABSTRACT FROM AUTHOR]

Group B Streptococcus (GBS) is the leading cause of neonatal disease worldwide, and invasive disease in adults is becoming more prevalent. Currently, some countries adopt an intrapartum antibiotic prophylaxis regime to help prevent the transmission of GBS from mother to neonate during delivery. This precaution has reduced the incidence of GBS-associated early-onset disease; however, rates of late-onset disease and stillbirths associated with GBS infections remain unchanged. GBS is still recognized as being universally susceptible to beta-lactam antibiotics; however, there have been reports of reduced susceptibility to beta-lactams, including penicillin, in some countries. Resistance to second-line antibiotics, such as erythromycin and clindamycin, remains high amongst GBS, with several countries noting increased resistance rates in recent years. Moreover, resistance to other antibiotic classes, such as fluoroquinolones and aminoglycosides, also continues to rise. In instances where patients are allergic to penicillin and second-line antibiotics are ineffective, vancomycin is administered. While vancomycin, a last resort antibiotic, still remains largely effective, there have been two documented cases of vancomycin resistance in GBS. This review provides a comprehensive analysis of the prevalence of antibiotic resistance in GBS and outlines the specific resistance mechanisms identified in GBS isolates to date. [ABSTRACT FROM AUTHOR]

Supershedding cattle shed Escherichia coli O157:H7 (O157) at ≥ 104 colony-forming units/g feces. We recently demonstrated that a supershed O157 (SS-O157) strain, SS-17, hyperadheres to the rectoanal junction (RAJ) squamous epithelial (RSE) cells which may contribute to SS-O157 persistence at this site in greater numbers, thereby increasing the fecal O157 load characterizing the supershedding phenomenon. In order to verify if this would be the signature adherence profile of any SS-O157, we tested additional SS-O157 isolates (n = 101; each from a different animal) in the RSE cell adherence assay. Similar to SS-17, all 101 SS-O157 exhibited aggregative adherence on RSE cells, with 56% attaching strongly (>10 bacteria/cell; hyperadherent) and 44% attaching moderately (1–10 bacteria/cells). Strain typing using Polymorphic Amplified Typing Sequences (PATS) analysis assigned the 101 SS-O157 into 5 major clades but not to any predominant genotype. Interestingly, 69% of SS-O157 isolates were identical to human O157 outbreak strains based on pulsed field gel electrophoresis profiles (CDC PulseNet Database), grouped into two clades by PATS distinguishing them from remaining SS-O157, and were hyperadherent on RSE cells. A subset of SS-O157 isolates (n = 53) representing different PATS and RSE cell adherence profiles were analyzed for antibiotic resistance (AR). Several SS-O157 (30/53) showed resistance to sulfisoxazole, and one isolate was resistant to both sulfisoxazole and tetracycline. Minimum inhibitory concentration (MIC) tests confirmed some of the resistance observed using the Kirby–Bauer disk diffusion test. Each SS-O157 isolate carried at least 10 genes associated with AR. However, genes directly associated with AR were rarely amplified: aac (3)-IV in 2 isolates, sul2 in 3 isolates, and tetB in one isolate. The integrase gene, int, linked with integron-based AR acquisition/transmission, was identified in 92% of SS-O157 isolates. Our results indicate that SS-O157 isolates could potentially persist longer at the bovine RAJ but exhibit limited resistance towards clinical antibiotics. [ABSTRACT FROM AUTHOR]

The reduction in childhood mortality noted in trials investigating azithromycin mass drug administration (MDA) for trachoma control has been confirmed by a recent large randomized controlled trial. Population-level implementation of azithromycin MDA may lead to selection of multiresistant pathogens. Evidence suggests that repeated azithromycin MDA may result in a sustained increase in macrolide and other antibiotic resistance in gut and respiratory bacteria. Current evidence comes from standard microbiological techniques in studies focused on a time-limited intervention, while MDA implemented for mortality benefits would likely repeatedly expose the population over a prolonged period and may require a different surveillance approach. Targeted short-term and long-term surveillance of resistance emergence to key antibiotics, especially those from the World Health Organization Access group, is needed throughout any implementation of azithromycin MDA, focusing on a genotypic approach to overcome the limitations of resistance surveillance in indicator bacteria. [ABSTRACT FROM AUTHOR]

In this Account, we describe our studies on the total synthesis of several natural products using intramolecular Nozaki–Hiyama–Takai–Kishi (NHTK) reactions. In each synthesis, an NHTK reaction is used to efficiently construct a medium-sized ring. These examples demonstrate the utility of the intramolecular NHTK reaction in natural product synthesis. 1 Introduction 2 Total Synthesis of (+)-Pestalotiopsin A 3 Total Synthesis of (+)-Cytosporolide A 4 Total Synthesis of (+)-Vibsanin A 5 Total Syntheses of (+)-Aquatolide and Related Humulanolides 6 Conclusion [ABSTRACT FROM AUTHOR]

The genetic basis of macrolide resistance was characterized in 59 Streptococcus pneumoniae isolates. All isolates were collected in 1995 and 1997 and were from invasive infections. The majority of the isolates (54 of 59 isolates) were erythromycin and clindamycin resistant (MLS(B)-phenotype) and carried the ermAM gene. Five isolates were erythromycin resistant but clindamycin susceptible (M-phenotype). Using PCR the mefE gene was detected in these five isolates. Contrary to the situation found in Canada and the USA, mefE-mediated erythromycin resistance in S. pneumoniae is uncommon in Belgium. [ABSTRACT FROM AUTHOR]

Background: Streptococcus mutans orchestrates the development of a biofilm that causes dental caries in the presence of dietary sucrose, and, in the bloodstream, S. mutans can cause systemic infections. The development of a cariogenic biofilm is dependent on the formation of an extracellular matrix rich in exopolysaccharides, which contains extracellular DNA (eDNA) and lipoteichoic acids (LTAs). While the exopolysaccharides are virulence markers, the involvement of genes linked to eDNA and LTAs metabolism in the pathogenicity of S. mutans remains unclear. Objective and Design: In this study, a parental strain S. mutans UA159 and derivative strains carrying single gene deletions were used to investigate the role of eDNA (ΔlytS and ΔlytT), LTA (ΔdltA and ΔdltD), and insoluble exopolysaccharides (ΔgtfB) in virulence in a rodent model of dental caries (rats) and a systemic infection model (Galleria mellonella larvae). Results: Fewer carious lesions were observed on smooth and sulcal surfaces of enamel and dentin of the rats infected with ∆lytS, ∆dltD, and ΔgtfB (vs. the parental strain). Moreover, strains carrying gene deletions prevented the killing of larvae (vs. the parental strain). Conclusions: Altogether, these findings indicate that inactivation of lytST and dltAD impaired S. mutans cariogenicity and virulence in vivo. [ABSTRACT FROM AUTHOR]

Shigella species remains a major diarrhoeagenic agent, affecting mostly children, with global high incidence and high mortality rate specially in developing areas. Although azithromycin is recommended for treatment of shigellosis, there are currently no CLSI susceptibility breakpoints, accordingly no routine antimicrobial susceptibility test is performed in the clinical laboratory. The purpose of this study was to estimate the prevalence, resistance profile and molecular epidemiology of azithromycin non-susceptible Shigella strains in Israel during a three year period. Shigella isolates (n = 1,170) referred to the National Reference Center during 2014–2016, were included in this study. Serotyping was performed by slide agglutination. Resistance genes, mph(A) and erm(B), were identified by PCR and the phenotype profile was determined by broth microdilution (BMD). Genetic relatedness was assessed by wgMLST. Decreased susceptibility to azithromycin (DSA) phenotype and genotype were detected in various Shigella species and serotypes related to diverse genetic backgrounds and antimicrobial profiles: 6% (26/423) of Shigella flexneri and 2% (16/747) of Shigella sonnei displayed DSA (MIC16 mg/L). Correlation of this phenotype with the presence of mph(A) and erm(B) genes was confirmed. All DSA-strains displayed resistance to ≥3 different antimicrobial classes. Among DSA-strains, 14% were resistant to quinolones and 5% displayed resistance to ceftriaxone. Most of these strains (32/42) were isolated from children in the southern and central regions of Israel. Clonality and significant relatedness was confirmed by PFGE and wgMLST. The presence of macrolide resistance genes among the different species and lineages reflects the transmissible nature of these genes. The emergence of DSA-Shigella reinforces the necessity to establish clinical breakpoints that would warrant routine testing, reporting and surveillance for this drug of choice. [ABSTRACT FROM AUTHOR]

Background: This study was conducted to investigate the genetic organization of erm-carrying Tn917 in clinical isolates of Streptococcus pyogenes. Tn917 is a nonconjugative transposon which is responsible for the spread of erythromycin resistance in bacterial isolates. Over the past two decades, erythromycin resistance rates have increased in S.pyogenes in many countries, To obtain information that may be useful in solving the spread of antimicrobial resistance, antibiotic-resistant genes can be identified as well as their association with mobile genetic elements. Material and Method: A total of 22 isolates of S. pyogenes isolates collected from the infection of upper respiratory system were examined, first by detecting their antibiotic susceptibility against three different antibiotics groups (Macrolides, Tetracycline, and lincosamides) then genomic DNA was extracted from each isolate for detection Tn917 by using specific primers to amplify erm(B) gene carried by this transposable element. Results: Results showed that there is a high level of resistance to erythromycin, (90.9%), then to Minocycline (68.1%), Lincomycin (59%), Tetracycline (54.5%), Clindamycin (50%), Azithromycin and Clarithromycin (36.3%), Doxycycline (31.8%) and then to Oxytetracycline (27.2%). Results also showed that five of S. pyogenes isolates were harboring Tn917 transposable element carrying erm gene. On the other hand, results showed that there are another isolates resistant to erythromycin that may possess a chromosomal or plasmid copy of the erythromycin resistance gene, or may the resistance caused by another structural erythromycin gene carried by other type of transposable elements rather than Tn917. Conclusion: Our findings suggest that the isolates of S. pyogenes are harboring chromosomal copy of Tn917 conferring erythromycin resistance. One possible explanation for the presence of genes at different isolates is due to erm gene, which was most likely located on Tn917. [ABSTRACT FROM AUTHOR]

Despite of the beneficial relevance of several lactic acid bacteria (LAB) in the food industry, micro‐organisms belonging to this group can determine spoilage in food products and carry a number of virulence and antibiotic resistance‐related genes. This study aimed on the characterization of beneficial and safety aspects of five bacteriocinogenic LAB strains (Lactobacillus curvatus 12—named L. curvatus UFV‐NPAC1), L. curvatus 36, Weissela viridescens 23, W. viridescens 31 and Lactococcus garvieae 36) isolated from an artisanal Brazilian calabresa, a traditional meat sausage. Regarding their beneficial aspects, all tested isolates were positive for mub, while EF226‐cbp, EF1249‐fbp and EF2380‐maz were detected in at least one tested strain; none of the isolates presented map, EFTu or prgB. However, evaluated strains presented a variable pattern of virulence‐related genes, but none of the strains presented gelE, cylA, efsA, cpd, int‐Tn or sprE. Moreover, other virulence‐related genes evaluated in this study were detected at different frequencies. L. curvatus 12 was generated positive results for ace, ccf, int, ermC, tetL, aac(6′)‐Ie‐aph(2″)‐Ia, aph(2″)‐Ib, aph(2″)‐Ic, bcrB, vanB and vanC2; L. curvatus 36: hyl, asa1, esp, int, ermC, tetK, aph(3′)‐IIIa, aph(2′′)‐Ic and vanC2; L. garvieae 32: asa1, ant(4′)‐Ia, aph(2′′)‐Ib, catA, vanA and vanC1; W. viridescens 23: esp, cob, ermB, aph(3′)‐IIIa, aph(2′′)‐Ic, vanA, vanB and vanC2; W. viridescens 31: hyl, esp, ermC, aph(3′)‐IIIa, aph(2′′)‐Ib, aph(2′′)‐Ic, catA, vanA and vanB. Despite presenting some beneficial aspects, the presence of virulence and antibiotic resistance genes jeopardize their utilization as starter or biopreservatives cultures in food products. Considering the inhibitory potential of these strains, an alternative would be the use of their bacteriocins as semi‐purified or pure technological preparation. Significance and Impact of the Study: The food industry has a particular interest in using bacteriocinogenic lactic acid bacteria (LAB) as starter, probiotics and/or biopreservatives in different food products. Characterization of additional beneficial features is important to identify new, multifunctional potential probiotic strains. However, these strains can only be applied in food products only after being properly characterized according their potential negative aspects, such as virulence and antibiotic resistance genes. A wide characterization of beneficial and safety aspects of bacteriocinogenic LAB is determinant to guide the proper utilization of these strains, or their purified bacteriocins, by the food industry. Significance and Impact of the Study: The food industry has a particular interest in using bacteriocinogenic lactic acid bacteria (LAB) as starter, probiotics and/or biopreservatives in different food products. Characterization of additional beneficial features is important to identify new, multifunctional potential probiotic strains. However, these strains can only be applied in food products only after being properly characterized according their potential negative aspects, such as virulence and antibiotic resistance genes. A wide characterization of beneficial and safety aspects of bacteriocinogenic LAB is determinant to guide the proper utilization of these strains, or their purified bacteriocins, by the food industry. [ABSTRACT FROM AUTHOR]

Aims: To determine the Staphylococcus aureus carriage rate in wild mammals in Aragon, northern Spain, to analyse their antimicrobial resistance phenotype/genotype and to characterize the recovered isolates. Methods and Results: Nasal and rectal swabs of 103 mammals were collected in Aragón during the period 2012–2015. Antimicrobial susceptibility, the presence of antimicrobial resistance genes and virulence factors were investigated. Molecular characterization was carried out by spa, MLST, agr and SCCmec. Staphylococcus aureus were recovered from 23 animals (22%). Four of the 23 S. aureus were methicillin‐resistant S. aureus (MRSA). Three MRSA were mecC‐positive and were isolated from European rabbits and were typed as t843 (ascribed to CC130). The remaining MRSA was a mecA‐carrying isolate from European hedgehog, typed as ST1‐t386‐SCCmecIVa‐agrIII and it harboured the blaZ, erm(C), ant(6)‐Ia and aph(3´)‐IIIa resistance genes. A high diversity of spa‐types was detected among the 19 methicillin‐susceptible S. aureus isolates, which showed high susceptibility to the antimicrobials tested. The tst gene and different combinations of staphylococcal enterotoxins were found. Conclusions: Staphylococcus aureus were detected in nasal and rectal samples of wild mammals. Wild rabbits could be a reservoir of mecC‐MRSA. Significance and Impact of the Study: This work provides information on the presence and characteristics of S. aureus from mammals in a defined geographic region in Spain. [ABSTRACT FROM AUTHOR]

The oral cavity contains a rich consortium of exopolysaccharide-producing microbes. These extracellular polysaccharides comprise a major component of the oral biofilm. Together with extracellular proteins, DNA, and lipids, they form the biofilm matrix, which contributes to bacterial colonization, biofilm formation and maintenance, and pathogenesis. While a number of oral microbes have been studied in detail with regard to biofilm formation and pathogenesis, the exopolysaccharides have been well characterized for only select organisms, namely Streptococcus mutans and Aggregatibacter actinomycetemcomitans. Studies on the exopolysaccharides of other oral organisms, however, are in their infancy. In this review, we present the current research on exopolysaccharides of oral microbes regarding their biosynthesis, regulation, contributions to biofilm formation and stability of the matrix, and immune evasion. In addition, insight into the role of exopolysaccharides in biofilms is highlighted through the evaluation of emerging techniques such as pH probing of biofilm colonies, solid-state nuclear magnetic resonance for macromolecular interactions within biofilms, and super-resolution microscopy analysis of biofilm development. Finally, exopolysaccharide as a potential nutrient source for species within a biofilm is discussed. [ABSTRACT FROM AUTHOR]

Bacteriocinogenic Enterococcus hirae ST57ACC and Pediococcus pentosaceus ST65ACC strains, previously isolated from artisanal cheese, were evaluated for their safety with the aim to determine whether they could be used as beneficial strains, especially in the control of Listeria monocytogenes. Both isolates survived simulated gastrointestinal conditions and showed high levels of auto- and co-aggregation with L. monocytogenes, although the hydrophobicity of cells varied. Using the agar-spot test with 33 commercial drugs from different groups, only anti-inflammatory drugs and drugs containing loratadine and propranolol hydrochloride were able to affect the growth of the tested strains. Both strains were resistant to 3 out of 11 antibiotics tested by the disc diffusion method, and low frequencies of antibiotic resistance-encoding genes were observed by PCR analysis. Tested strains neither presented biogenic amine-related genes nor produced these substances. Aside from some antibiotic resistance characteristics, the tested strains were considered safe as they lack other virulence-related genes. E. hirae ST57ACC and P. pentosaceus ST65ACC both presented beneficial properties, particularly their ability to survive gastrointestinal conditions and to aggregate with L. monocytogenes, which can facilitate the elimination of this pathogen. Further studies should be conducted to better understand these interactions. [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the probiotic properties of Lactobacillus casei and Lactobacillus fermentum strains, as well as to select novel and safe strains for future development of functional fermented products. The in vitro auto-aggregation, co-aggregation, hydrophobicity, β-galactosidase production, survival to gastrointestinal tract (GIT), and antibiotic susceptibility were evaluated. The selected strains were additionally tested by the presence of genes encoding adhesion, aggregation and colonization, virulence factors, antibiotic resistance, and biogenic amine production, followed by the evaluation of acidifying kinetic parameters in milk, and survival of the strains under simulated GIT conditions during refrigerated storage of fermented milk. Most strains of both species showed high auto-aggregation; some strains showed co-aggregation ability with other lactic acid bacteria (LAB) and/or pathogens, and both species showed low hydrophobicity values. Seven L. casei and six L. fermentum strains produced β-galactosidase enzymes, and ten strains survived well the simulation of the GIT stressful conditions evaluated in vitro. All strains were resistant to vancomycin, and almost all the strains were resistant to kanamycin. L. casei SJRP38 and L. fermentum SJRP43 were distinguished among the other LAB strains by their higher probiotic potential. L. fermentum SJRP43 presented fewer genes related to virulence factors and antibiotic resistance and needed more time to reach the maximum acidification rate (Vmax). The other kinetic parameters were similar. Both strains survived well (> 8 log10 CFU/mL) to the GIT-simulated conditions when incorporated in fermented milk. Therefore, these strains presented promising properties for further applications in fermented functional products. [ABSTRACT FROM AUTHOR]

The cell wall‐anchored protein‐encoding spj gene on staphylococcal cassette chromosome mec IVl (SCCmecIVl) was found to vary in size because of its 22‐ and 86‐aa repeat domains. The 22‐aa repeats are the more flexible of the two repeats, comprising three 11‐aa units, and were classified into three groups with eleven types. The 11/22‐aa repeats are longer in individuals with bullous impetigo, shorter in those with invasive disease and were absent in a fatal case, this last one having been rapidly diagnosed by PCR. IS431‐flanking pUB110 (bleO, aadD) is present on SCCmecIVl at 90%. The bacterial surface has the spj product and a unique surface layer. [ABSTRACT FROM AUTHOR]

Introduction: Enterococci commonly inhabit the gastrointestinal tract of both human and animals; however, they have emerged as a leading cause of several infections with substantial morbidity and mortality. Their ability to acquire resistance combined with intrinsic resistance to various antimicrobials makes treatment of enterococcal infections challenging. Materials and methods: The aim of the study was to evaluate the antimicrobial resistance pattern, and assess the prevalence of multidrug resistance (MDR) and extensive drug resistance (XDR) among enterococcal isolates, collected from different clinical sources, in Mansoura University Hospitals, Egypt. Results: Antibiotic sensitivity testing revealed elevated levels of resistance among enterococcal clinical isolates (N=103). All E. faecium (N=32) and 74.6% of E. faecalis isolates (N=71) were MDR, while two E. faecalis and four E. faecium isolates were XDR. High level gentamicin resistance was detected in 79.6%, most of them carried the aac(6')-Ie-aph(2")-Ia gene. High level streptomycin resistance was seen in 36.9%, of which 52.6% carried the ant(6')-Ia gene. Resistance to macrolides and lincosamides were mediated by ermB (92.2%) and msrA/B (42.7%). tetK, tetL, and tetM genes were detected among tetracyclines resistant isolates. Resistance to vancomycin was detected in 15.5%, where vanB and vanC1 gene clusters were detected in VRE isolates. Ten isolates (9.7%) were resistant to linezolid, eight of which harbored the optrA gene. Vancomycin and linezolid resistant enterococci were more likely to exhibit strong/moderate biofilm formation than vancomycin and linezolid sensitive ones. Conclusion: Elevated levels of resistance to different classes of antimicrobial agents and emergence of MDR and XDR strains pose a major threat with limited therapeutic options for infections caused by this emerging pathogen. [ABSTRACT FROM AUTHOR]

Transferable genetic elements conferring macrolide resistance in Streptococcus pneumoniae can encode the efflux pump and ribosomal protection protein, mef (E)/ mel , in an operon of the macrolide efflux genetic assembly (Mega) element- or induce ribosomal methylation through a methyltransferase encoded by erm (B). During the past 30 years, strains that contain Mega or erm (B) or both elements on Tn 2010 and other Tn 916 -like composite mobile genetic elements have emerged and expanded globally. In this study, we identify and define pneumococcal isolates with unusually high-level macrolide resistance (MICs > 16 μg/ml) due to the presence of the Mega element [ mef (E)/ mel ] alone. High-level resistance due to mef (E)/ mel was associated with at least two specific genomic insertions of the Mega element, designated Mega-2.IVa and Mega-2.IVc. Genome analyses revealed that these strains do not possess erm (B) or known ribosomal mutations. Deletion of mef (E)/ mel in these isolates eliminated macrolide resistance. We also found that Mef(E) and Mel of Tn 2010 -containing pneumococci were functional but the high-level of macrolide resistance was due to Erm(B). Using in vitro competition experiments in the presence of macrolides, high-level macrolide-resistant S. pneumoniae conferred by either Mega-2.IVa or erm (B), had a growth fitness advantage over the lower-level, mef (E)/ mel -mediated macrolide-resistant S. pneumoniae phenotypes. These data indicate the ability of S. pneumoniae to generate high-level macrolide resistance by macrolide efflux/ribosomal protection [Mef(E)/Mel] and that high-level resistance regardless of mechanism provides a fitness advantage in the presence of macrolides. [ABSTRACT FROM AUTHOR]

Background: Streptococcus gallolyticus subspecies (subsp.) pasteurianus, previously known as Streptococcus bovis biotype II/2, has been described as a causative agent of endocarditis, neonatal sepsis, meningitis, bacteremia, and colorectal carcinoma in humans. The aim of this study was to characterize the erythromycin and tetracycline resistance genes of S. gallolyticus subsp. pasteurianus strains isolated from patients with septicemia and bacteremia in Thailand. Methods: The clinical isolates of Streptococcus gallolyticus were identified by using conventional biochemical tests, PCR, and sodA gene sequence analysis. The erythromycin and tetracycline susceptibilities were determined by disk diffusion and agar dilution methods, while the resistance genes were identified by nucleotide sequence analysis. Results: From a total of 108 blood cultures, 36 (33%) were identified as S. gallolyticus subsp. pasteurianus with the nucleotide sequence identities of partial sodA gene with the reference strains ranging from 98.1 to 100%. Of these, 25 (69.4%) contained erythromycin resistance genes and erm(B) was the most predominant gene (30.6%), followed by erm(T) (19.4%) and mef(A) (5.6%). In addition, erm(B) was also detected in combination with lnu(B) (8.3%), erm(T) and mef(A) (2.8%), and mef(A) and lnu(B) (2.8%). It was interesting to note that lnu(B) was detected for the first time in S. gallolyticus subsp. pasteurianus in this study. For tetracycline resistance genes, tet(L) and tet(M) were detected at 13.9% and 11.1%, respectively. However, tet(M) in combination with tet(L) was detected most commonly at 69.4% and with tet(L) and tet(O) at 5.6%. Conclusions: A number of erythromycin and tetracycline resistance genes were detected in S. gallolyticus subsp. pasteurianus strains circulating in Thailand. [ABSTRACT FROM AUTHOR]

Background: Streptococcus pneumoniae (S. pneumoniae) is one of the most common causes of human diseases in young children. Macrolides are commonly antibiotics used for empirical treatment of community-acquired respiratory infections. The purpose of this study was to determine antibiotic resistance pattern as well as the relationship between macrolide resistance and the major mechanisms of resistance in pneumococci isolated from healthy children. Materials and Methods: In this cross-sectional study, 43 isolates of S. pneumoniae were collected from healthy children in Ardabil. Resistance pattern against tested antibiotics was determined using the disk diffusion method. The Minimum Inhibitory Concentration (MIC) of erythromycin was determined using the E-test strips. The mefA/E and ermB gene were detected in erythromycin-resistant isolates using the specific primers and Polymerase Chain Reaction (PCR) technique. Results: According to antimicrobial susceptibility testing, 74.4% of the isolates were resistant to erythromycin, 95.3% to penicillin, 81.3% to co-trimoxazole, 72% to azithromycin, 41.8% to tetracycline, 27.9% to clindamycin, and 16.2% to chloramphenicol. All isolates were susceptible to levofloxacin and vancomycin. In the case of rifampin, 95.3% of the isolates were sensitive and 4.6% semisensitive. The MIC of erythromycin for resistant isolates was between 1.5 and ≥ 256 μg/ml. PCR results revealed that 100% of erythromycin-resistant isolates contained mefA/E gene and 81.25% contained both the ermB and mefA/E genes. Conclusion: The prevalence of antibiotic-resistant strains of S. pneumoniae, especially resistance to macrolides, was high among healthy children in Ardabil. According to the results of this study, we suggest using levofloxacin, rifampin and vancomycin antibiotics as an appropriate prophylactic regimen in pneumococcal infections. [ABSTRACT FROM AUTHOR]

Aim: The aim of this study is to assess the variations in the levels of Streptococcus mutans in saliva before and after short-term daily intake of the probiotic lozenge. To estimate the prevalence of S. mutans serotypes e, f, and k and its variations in salivary levels before and after short-term daily intake of the probiotic lozenge.Materials and Methods: This double-blind randomized controlled trial was conducted among 60 children of 6-12 years old, who were given lozenge twice daily containing probiotic bacteria twice daily, one in the morning and another in the evening after brushing for 1 month. The placebo lozenge group also followed the same protocol. A pre- and post-quantitative analysis of S. mutans and serotypes e, f, and k level were done in the saliva of both experiment and control groups.Statistical Analysis: The data thus collected were statistically evaluated using repeated measures of ANOVA, followed by post hoc test and independent t-test.Results and Conclusion: When compared with baseline, a significant reduction of S. mutans was seen after 1 month intervention and 6 months follow-up. The prevalence of serotype e was found to be 5% and serotype k was found to be 12%, whereas none of the samples carried serotype f. Future long-term research with a larger sample size is required to determine the prevalence of S. mutans serotype and longer evaluation period is required to assess the preventive role against caries development. [ABSTRACT FROM AUTHOR]

According to recent studies, the importance of MLS (macrolide-lincosamide-streptogramin) resistance phenotypes and genes in enterococci are reflected in the fact that they represent reservoirs of MLS resistance genes. The aim of this study was to investigate distribution of MLS resistance genes and phenotypes in community- and hospital-acquired enterococcal isolates and to determine their prevalence. The MLS resistance phenotypes (cMLSb, iMLSb, M/MSb, and L/LSa) were determined in 245 enterococcal isolates were characterized using the double-disc diffusion method. Specific primers were chosen from database sequences for detection of the MLS resistance genes (ermA, ermB, ermC, msrA/B, lnuA, lnuB, and lsaA) in 60 isolates of enterococci by end-point PCR. There was no linezolid-resistant enterococcal isolate. Only one vancomycin-resistant (0.6%) isolate was found and it occurred in a community-acquired enterococcal isolate. The most frequent MLS resistance phenotype among enterococcal isolates was cMLSb (79.7% community- and 67.9% hospital-acquired). The most common identified MLS resistance genes among enterococcal isolates were lsaA (52.9% community- and 33.3% hospital-acquired) and ermB (17.6% community- and 33.3% hospital-acquired). The most prevalent MLS gene combination was lnuA + lsaA (five enterococcal isolates). The ermB gene encoded cMLSb phenotype, and it was identified in only one isolate that displayed iMLSb resistance phenotype. Based on the results obtained, we can conclude that the most frequent MLS resistance phenotype among enterococcal isolates was cMLSb. Surprisingly, a vancomycin-resistant enterococcal isolate was identified in a community-acquired enterococcal isolate. This study shows that enterococci may represent a major reservoir of ermB, lsaA, and lnuA genes., (© 2022. The Author(s), under exclusive licence to the Japan Antibiotics Research Association.)

Background Frequent use of antibiotics is thought to create selection pressure by clearing susceptible bacteria and allowing resistant bacteria to spread in a community. A cluster-randomized trial comparing 2 different frequencies of mass azithromycin distributions for trachoma provided a convenient experiment for determining the causal relationship between antibiotic consumption and antibiotic resistance. Methods Twenty-four communities were randomized to either annual or biannual mass azithromycin distributions for trachoma. Randomization was stratified on health catchment area and trachoma prevalence. Swabs were processed for the genetic macrolide resistance determinants ermB and mefA/E in a masked fashion from a random sample of 120 preschool children before treatment and another 120 children after 2 years of mass antibiotics. Results Macrolide resistance determinants were similar in the 12 annually and 12 biannually treated communities before treatment, with a median prevalence among preschool children of 20% (interquartile range [IQR], 10%–40%) in each group. By 24 months, macrolide resistance determinants were found more commonly in the biannually treated communities (median, 60% [IQR, 50%–80%]) than the annually treated communities (median, 40% [IQR, 20%–40%]; P <.001). Adjusting for baseline, the 24-month prevalence of macrolide resistance determinants in the biannual group was 29.4% higher than that of the annual group (95% confidence interval, 10.5%–56.7%). Conclusions This randomized trial used direct genetic methods to confirm the causal relationship of community antibiotic consumption and antibiotic resistance. Communities randomized to less frequent use of antibiotics had a significantly lower prevalence of genetic antibiotic resistance determinants. Clinical Trials Registration NCT00792922. [ABSTRACT FROM AUTHOR]

Lactobacillus fermentum colonizing gastrointestinal and urogenital tracts of humans and animals is widely used in manufacturing of fermented products and as probiotics. These bacteria may function as vehicles of antibiotic resistance genes, which can be transferred to pathogenic bacteria. Therefore, monitoring and control of transmissible antibiotic resistance determinants in these microorganisms is necessary to approve their safety status. The aim of this study was to characterize erythromycin and tetracycline resistance of L. fermentum isolates and to estimate the potential transfer of resistance genes from lactobacilli to the other Gram-positive and Gram-negative bacteria. Among six L. fermentum strains isolated from human feces and commercial dairy products, five strains demonstrated phenotypic resistance to tetracycline. PCR screening for antibiotic resistance determinants revealed plasmid-located tetracycline resistance genes tet(K) and tet(M) in all strains and erythromycin resistance genes erm(B) in the chromosome of L. fermentum 5-1 and erm(C) in the plasmid of L. fermentum 3-4. All tested lactobacilli lacked conjugative transposon Tn916 and were not able to transfer tetracycline resistance genes to Staphylococcus aureus, Staphylococcus epidermidis, Listeria monocytogenes, Acinetobacter baumannii, Citrobacter freundii, and Escherichia coli by filter mating. Staphylococcus haemolyticus did not accept erythromycin resistance genes from corresponding Lactobacillus strains. Thus, in the present study, L. fermentum was not implicated in the spread of erythromycin and tetracycline resistance, but still these strains pose the threat to the environment and human health because they harbored erythromycin and tetracycline resistance genes in their plasmids and therefore should not be used in foods and probiotics. [ABSTRACT FROM AUTHOR]

The environment, human, and animals play an important role in the spread of antibiotic-resistant bacteria. Enterococci are members of the gastrointestinal tracts of humans and animals and represent important reservoirs of antibiotic resistance genes. Until today, few studies have examined antibiotic susceptibility in enterococci isolated from primates. Therefore, the present study investigated species distribution, antibiotic susceptibility, and resistance genes in enterococci isolated from wild and captive black capuchins monkeys (Sapajus nigritus) in Rio Grande do Sul, South Brazil. A total of 24 swabs/fecal samples were collected, including 19 from wild monkeys living in two forest fragments [São Sebastião do Caí (SSC) and Santa Cruz do Sul (SCS)], and five in captive [Parque Zoológico da Fundação Zoobotânica (ZOO)], between August 2016 and November 2017. Fifteen colonies were randomly selected from each sample. Enterococci were identified by MALDI-TOF, tested for susceptibility to 12 antibiotics; and screened for tet (S), tet (M), tet (L), msrC , and erm (B) genes by PCR. Two-hundred ninety-six enterococci were isolated (SSC n = 137; SCS n = 86; ZOO n = 73) and differences in Enterococcus species distribution were detected on three monkey groups, with low abundance in SCS (1 - D = 0.2), followed by ZOO (1 - D = 0.68), and SSC (1 - D = 0.73). The enterococci frequently recovered include the following: Enterococcus faecalis (42.6%), E. hirae (29.1%), and E. faecium (15.9%). Antibiotic-nonsusceptible was observed in 202 (67.9%) strains. The rate of non-susceptibility to rifampicin, tetracycline, erythromycin, nitrofurantoin, chloramphenicol, and ampicillin was 46%, 26%, 22% and 19%, 13%, 0.3%, and 0.3%, respectively. All strains were susceptible to vancomycin, streptomycin, gentamycin, and linezolid. Forty-three (14.52%) isolates were identified as multidrug resistant (MDR), and the highest number of MDR enterococci were E. faecium recovered from wild monkeys living close to a hospital and water treatment plant. Elevated rates of antibiotic resistance genes msr C and tet (L) were isolates from ZOO. In conclusion, differences in the frequency of enterococci species, antibiotic-nonsusceptible and antibiotic resistance genes in all groups of monkeys were identified. These data suggest that anthropogenic activities could have an impact in the resistome of primate gut enterococci communities. [ABSTRACT FROM AUTHOR]

Introduction: The emergence of erythromycin resistant Gram positive cocci in various parts of the world has become worrisome in clinical settings in recent times, however there is little or no information on the determinants of erythromycin resistance from Nigeria. Aim: This study investigated the determinants of erythromycin resistance in clinical Gram positive cocci bacteria isolated from Nigerian patients. Materials and Methods: Assembly of isolates of non-duplicate staphylococci from various clinical specimens from south western and northern hospitals of Nigeria were collected. While enterococci were obtained from our culture collection which were previously collected from healthy people from community. Characterisation of 25 staphylococci and enterococci each were done using standard microbiology procedures, susceptibility pattern to erythromycin and other panel of antimicrobial agents including Minimum Inhibitory Concentrations (MICs) to erythromycin was determined. Erythromycin resistance genes were amplified using Polymerase Chain Reaction (PCR). Results: None of the strains had ermA and mefA but the strains showed heterogeneous possession of ermB, ermC, msrA and msrB genes in no particular pattern including multiple gene acquisition. MIC50 and MIC90 of staphylococci strains to erythromycin were 2 μg/mL and >64 μg/mL, respectively; wherein 7 (28%) were sensitive to erythromycin, while 11 (44%) of enterococci were sensitive to erythromycin with MIC50 and MIC90 of 1 and >64 μg/mL, respectively. One of the staphylococci isolates had inducible clindamycin resistance. Conclusion: In conclusion, high level staphylococci and enterococci resistance was found to various antibiotics with limited therapeutic option. Ribosomal methylation and efflux are the main resistant determinants found in these isolates. [ABSTRACT FROM AUTHOR]

Antibiotics resistant genes (ARGs) are frequently detected in different media, such as wastewater, sewage sludge, and livestock manure. ARGs have been evidenced to have more and more threats to our environment because of their increase in species and total abundances causing more attention especially in horizontal gene transfer (HGT). The integron, an important form of mobile gene elements transfer ARGs through HGT, is demonstrated to have a high risk to human beings, and the class 1 integron (intl1), a predominant integron, is a marker of the process of horizontal gene transfer. Tetracycline, sulfonamides, macrolide, beta-lactam, trimethoprim, and quinolone plasmid-mediated resistance genes, which are frequently detected in various media and different treatment processes, have been reported to have significant correlations with intl1. Considering the risk of ARGs, especially those associated with intl1, the removal of intl1 and associated ARGs from water, sludge and livestock manure has attracted much more attention recently. Therefore, the mechanism and removal of intl1 and associated ARGs in water depth treatment, sludge and livestock manure digestion and composting process are reviewed in this paper. Besides, the limitation of the current study in this field is discussed, and the key points in the future investigation are provided. [ABSTRACT FROM AUTHOR]

Doxycycline is an antibiotic that, in addition to the classic antibacterial use, is also prescribed to fight parasitic diseases, like heartworm disease in dogs. Despite the concern that the overuse of this antibiotic may decrease susceptibility of clinically important bacteria, the consequences of the prolonged doxycycline therapy in heartworm-infected dogs have never been studied before. We have analyzed the impact of this therapy on Staphylococcus aureus and Enterococcus antimicrobial resistance. In this study, 17 heartworm-infected dogs (10 that had completed the doxycycline treatment and 7 dogs that had not yet begun) were included. Twenty-four isolates of Staphylococcus aureus were obtained from two locations of each dog. After treatment, 73.3% of isolates were resistant to at least one antibiotic but only 22.2% of isolates before treatment. Most of doxycycline resistant isolates were obtained from dogs that have received treatment. Erythromycin resistance or intermediate susceptibility was detected in 45.6% of isolates, most of them from dogs after treatment. For Enterococci, 48 isolates were obtained from fecal samples (25 before treatment and 23 after treatment). Before treatment, 32% of isolates were resistant at least to one antibiotic while after, this data increase up to 65%. Comparing isolates before and after treatment, a clear increase in resistance to doxycycline (12% against 21.74%) and erythromycin (20% against 39.13%) was observed. Although the present work is a preliminary research, the results encourages the development of further studies to determinate the effect of prolonged doxycycline therapy on antimicrobial resistance. [ABSTRACT FROM AUTHOR]

The mef (A) gene was originally identified as the resistance determinant responsible for type M resistance to macrolides, a phenotype frequently found in clinical isolates of Streptococcus pneumoniae and Streptococcus pyogenes. MefA was defined as a secondary transporter of the major facilitator superfamily driven by proton-motive force. However, when characterizing the mef (A)-carrying elements Tn 1207.1 and Φ1207.3, another macrolide resistance gene, msr (D), was found adjacent to mef (A). To define the respective contribution of mef (A) and msr (D) to macrolide resistance, three isogenic deletion mutants were constructed by transformation of a S. pneumoniae strain carrying Φ1207.3: (i) Δ mef (A)–Δ msr (D); (ii) Δ mef (A)– msr (D); and (iii) mef (A)–Δ msr (D). Susceptibility testing of mutants clearly showed that msr (D) is required for macrolide resistance, while deletion of mef (A) produced only a twofold reduction in the minimal inhibitory concentration (MIC) for erythromycin. The contribution of msr (D) to macrolide resistance was also studied in S. pyogenes , which is the original host of Φ1207.3. Two isogenic strains of S. pyogenes were constructed: (i) FR156, carrying Φ1207.3, and (ii) FR155, carrying Φ1207.3/Δ msr (D). FR155 was susceptible to erythromycin, whereas FR156 was resistant, with an MIC value of 8 μg/ml. Complementation experiments showed that reintroduction of the msr (D) gene could restore macrolide resistance in Δ msr (D) mutants. Radiolabeled erythromycin was retained by strains lacking msr (D), while msr (D)-carrying strains showed erythromycin efflux. Deletion of mef (A) did not affect erythromycin efflux. This data suggest that type M resistance to macrolides in streptococci is due to an efflux transport system of the ATP-binding cassette (ABC) superfamily, in which mef (A) encodes the transmembrane channel, and msr (D) the two ATP-binding domains. [ABSTRACT FROM AUTHOR]

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Abstract: The aim of this study was to evaluate the antibiotic susceptibility of lactic acid bacteria (LAB) and coagulase‐negative staphylococci (CNS) strains isolated from naturally fermented smoked pork produced in Hunan, China. A total of 48 strains were isolated by selective medium and identified at the species level by 16S rRNA gene sequencing as follows: Staphylococcus carnosus (23), Lactobacillus plantarum (12), Lactobacillus brevis (10), Lactobacillus sakei (1), Weissella confusa (1), and Weissella cibaria (1). All strains were typed by RAPD‐PCR, and their susceptibility to 15 antibiotics was determined and expressed as the minimum inhibitory concentration (MIC) using agar dilution method. High resistance to penicillin G, streptomycin, gentamycin, vancomycin, chloramphenicol, norfloxacin, ciprofloxacin, kanamycin, and neomycin was found among the isolates. All the strains were sensitive to ampicillin, while the susceptibility to tetracycline, oxytetracycline, erythromycin, lincomycin, and roxithromycin varied. The presence of relevant resistance genes was investigated by PCR and sequencing, with the following genes detected: str(A), str(B), tet(O), tet(M), ere(A), and catA. Eleven strains, including 3 S. carnosus, 6 L. plantarum, and 2 L. brevis, harbored more than 3 antibiotic resistance genes. Overall, multiple antibiotic resistance patterns were widely observed in LAB and S. carnosus strains isolated from Hunan smoked pork. Risk assessment should be carried out with regard to the safe use of LAB and CNS in food production. Practical Application: We evaluated the antibiotic resistance of lactic acid bacteria and coagulase‐negative staphylococci strains isolated from Chinese naturally fermented smoked pork. Our results may provide important data on establishing breakpoint standards for LAB and CNS and evaluating the safety risk of these strains for commercial use. [ABSTRACT FROM AUTHOR]

Humic and fulvic acids constitute humic substances, a complex mixture of many different acids containing carboxyl and phenolate groups, which are not only the principal soil fertility factors but also the main pollutants present in landfill leachates or natural organic matter in water. Due to their low bacterial biodegradability, fungal biodegradation processes are key for their removal. The present study compiles and comments all the available literature on decomposition of aqueous humic substances by fungi or by their extracellular enzymes alone, focusing on the influence of the reaction conditions. The biodegradation extent mainly depends on the characteristics and concentration of the humic compounds, the type of microorganisms selected, the inoculation mode, the C and N sources, the presence of certain chemicals in the medium, the availability of oxygen, the temperature, and the pH. [ABSTRACT FROM AUTHOR]

A biofilm is an aggregate of microorganisms in which cells adhere to biological or non- biological surfaces and is responsible for various infectious diseases. Infections caused by Staphylococcus aureus, including pneumonia, endocarditis, and osteomyelitis, are often associated with colonization and biofilm formation. Although lipoteichoic acid (LTA) is involved in biofilm formation, the specific role of LTA is not clearly understood. In this study, we demonstrated that LTA released from Lactobacillus plantarum could inhibit S. aureus biofilm formation and aggregation without affecting the growth of S. aureus in various in vitro and in vivo models. L. plantarum LTA (Lp.LTA) also inhibited biofilm formation of S. aureus clinical isolates, including a methicillin-resistant strain. Remarkably, Lp.LTA not only interfered with S. aureus biofilm formation, but it also disrupted a pre-formed biofilm. Mechanism studies demonstrated that Lp.LTA inhibited expression of the ica-operon, which is responsible for the production of poly-N-acetylglucosamine, a key molecule required for S. aureus biofilm development. Lp.LTA increased the release of autoinducer-2 from S. aureus, which contributed to the inhibition of S. aureus biofilm formation. Moreover, Lp.LTA treatment enhanced susceptibility of the biofilm to various antibiotics and to macrophages. Interestingly, Lp.LTA without D-alanine moieties was not able to inhibit biofilm formation by S. aureus. In conclusion, the present study suggests that LTA can inhibit S. aureus biofilm formation, and therefore could be applied for preventing and/or treating infectious diseases caused by S. aureus biofilms. [ABSTRACT FROM AUTHOR]

Dental caries is a biofilm-dependent oral disease and Streptococcus mutans is the known primary etiologic agent of dental caries that initiates biofilm formation on tooth surfaces. Although some Lactobacillus strains inhibit biofilm formation of oral pathogenic bacteria, the molecular mechanisms by which lactobacilli inhibit bacterial biofilm formation are not clearly understood. In this study, we demonstrated that Lactobacillus plantarum lipoteichoic acid (Lp.LTA) inhibited the biofilm formation of S. mutans on polystyrene plates, hydroxyapatite discs, and dentin slices without affecting the bacterial growth. Lp.LTA interferes with sucrose decomposition of S. mutans required for the production of exopolysaccharide, which is a main component of biofilm. Lp.LTA also attenuated the biding of fluorescein isothiocyanate-conjugated dextran to S. mutans, which is known to have a high affinity to exopolysaccharide on S. mutans. Dealanylated Lp.LTA did not inhibit biofilm formation of S. mutans implying that D-alanine moieties in the Lp.LTA structure were crucial for inhibition. Collectively, these results suggest that Lp.LTA attenuates S. mutans biofilm formation and could be used to develop effective anticaries agents. [ABSTRACT FROM AUTHOR]

Abstract: A selective, sensitive and rapid liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the determination of tigecycline (TGC) in human plasma, using tigecycline‐d9 as an internal standard (IS). Analytical samples were prepared using a protein precipitation method coupled with a concentration process. The analyte and IS were separated on a reversed‐phase Waters Acquity UPLC® BEH‐C18 column (2.1 × 50 mm i.d., 1.7 μm) with a flow rate of 0.25 mL/min. The mobile phase consisted of water, containing 0.2% formic acid (v/v) with 10 m m ammonium formate (A) and acetonitrile (B). The mass spectrometer was operated in selected reaction monitoring mode through electrospray ionization ion mode using the transitions of m/z 586.2 → 513.1 and m/z 595.1 → 514.0 for TGC and IS, respectively. The linearity of the method was in the range of 10–5000 ng/mL. Intra‐ and inter‐batch precision (CV) for TGC was <9.27%, and the accuracy ranged from 90.06 to 107.13%. This method was successfully applied to the analysis of samples from hospital‐acquired pneumonia patients treated with TGC, and a validated population pharmacokinetic model was established. This developed method could be useful to predict pharmacokinetics parameters and valuable for further pharmacokinetics/pharmacodynamics studies. [ABSTRACT FROM AUTHOR]

Aims The aim of this study was to determine the intestinal microbiota of pirarucu ( Arapaima gigas) in different growth stages (adult and fingerlings) and to isolate and identify potential probiotic bacteria. Methods and Results High-throughput sequencing analysis of the intestinal contents revealed that the majority of sequences belonged to the Proteobacteria, Fusobacteria and Firmicutes phyla. At the genus level, the greatest number of sequences belonged to Bradyrhizobium in adult fish, while Cetobacterium was the most abundant in juvenile fish. Twenty-three lactic-acid bacteria ( LABs) were isolated on MRS agar from healthy juvenile fish. The isolates were tested in vitro for probiotic properties. Two isolates (identified as strains of Lactococcus lactis subsp. lactis and Enterococcus faecium) displayed antagonism against all 10 pathogens tested, were nonhaemolytic and maintained good viability for at least 3 weeks when supplemented to fish diets. The presence of a number of antibiotic resistance genes ( ARGs), conferring resistance to erythromycin, tetracycline and chloramphenicol, was investigated by PCR. Conclusions The absence of ARGs investigated the potential to antagonize pathogens, and favourable growth and survival characteristics indicate that these autochthonous isolates have the potential to be considered probiotics, which will be studied in future in vivo experiments. Significance and Impact of the Study This study has demonstrated, for the first time, the normal microbiota in the A. gigas intestine during different life stages and the presence of LAB strains. It also demonstrated LAB antibiotic resistance and antagonistic behaviour against pathogens isolated from the same fish. [ABSTRACT FROM AUTHOR]

Macrolides represent a large family of protein synthesis inhibitors of great clinical interest due to their applicability to human medicine. Macrolides are composed of a macrocyclic lactone of different ring sizes, to which one or more deoxy-sugar or amino sugar residues are attached. Macrolides act as antibiotics by binding to bacterial 50S ribosomal subunit and interfering with protein synthesis. The high affinity of macrolides for bacterial ribosomes, together with the highly conserved structure of ribosomes across virtually all of the bacterial species, is consistent with their broad-spectrum activity. Since the discovery of the progenitor macrolide, erythromycin, in 1950, many derivatives have been synthesised, leading to compounds with better bioavailability and acid stability and improved pharmacokinetics. These efforts led to the second generation of macrolides, including well-known members such as azithromycin and clarithromycin. Subsequently, in order to address increasing antibiotic resistance, a third generation of macrolides displaying improved activity against many macrolide resistant strains was developed. However, these improvements were accompanied with serious side effects, leading to disappointment and causing many researchers to stop working on macrolide derivatives, assuming that this procedure had reached the end. In contrast, a recent published breakthrough introduced a new chemical platform for synthesis and discovery of a wide range of diverse macrolide antibiotics. This chemical synthesis revolution, in combination with reduction in the side effects, namely, 'Ketek effects', has led to a macrolide renaissance, increasing the hope for novel and safe therapeutic agents to combat serious human infectious diseases. [ABSTRACT FROM AUTHOR]

Background: Despite the employment of sanitary regimes, contact transmission of the aetiological agents of hospital infections is still exceedingly common. The issue of microbe transmission becomes particularly important when facing multidrug-resistant microorganisms such as methicillin-resistant staphylococci. In the case of deficiencies in cleaning and disinfection procedures, hospital equipment made of copper alloys can play an important role, complementing traditional hospital hygiene procedures. The objective of this study was to characterize staphylococcal strains isolated from touch surfaces in Polish hospital wards in terms of their drug resistance, ability to form biofilm and susceptibility to antimicrobial activity of copper alloys. Methods: The materials for the study were 95 staphylococcal strains isolated from touch surfaces in 13 different hospital wards from Małopolska province (the south of Poland). Phenotypic and genotypic antibiotic resistance were checked for erythromycin, clindamycin, gentamycin, ciprofloxacin, trimethoprim/sulfamethoxazole and mupirocin. Biofilm formation ability for the tested strains was checked with the use of culture on Congo red agar. Susceptibility to copper, tin bronze, brass and new silver was tested using a modification of the Japanese standard. Results: Over 67% of the analysed staphylococcal strains were methicillin-resistant (MR). Four strains were resistant to all of the tested antibiotics, and 14 were resistant to all except mupirocin. Strains classified as MR had significantly increased resistance to the remaining antibiotic groups. About one-third of the analysed strains revealed biofilm-forming ability. Among the majority of species, biofilm-forming and non-biofilm-forming strains were distributed evenly; in the case of S. haemolyticus only, negative strains accounted for 92.8%. Susceptibility to copper alloys was different between strains and rather lower than in the case of the SA strain selected for comparison. Conclusions: Coagulase-negative staphylococci, the most commonly isolated in Polish hospital wards, should not be neglected as an infection risk factor due their high antibiotic resistance. Our experiments confirmed that touch surfaces made of copper alloys may play an important role in eliminating bacteria from the hospital environment. [ABSTRACT FROM AUTHOR]

In this study, prevalence, biotechnological and safety profiles of 588 Enterococcus isolates isolated from raw milk and Istrian cheese during different stages of ripening were analyzed. Despite the low and variable presence of enterococci in milk ((3.65±2.93) log CFU/mL), highly comparable enterococcal populations were established after 30 days of cheese ripening ((7.96±0.80) log CFU/g), confirming Enterococcus spp. as a major part of the core microbiota of Istrian cheese. The dominant species were E. faecium (53.8 %) and E. faecalis (42.4 %), while minor groups, consisting of E. durans (2.84 %) and E. casseliflavus (0.95 %), also occurred. A pronounced intraspecies variability was noticed based on molecular fingerprinting, with 35 strains (genotypes) detected. Most of the genotypes were farm-specific with one third being shared between the farms. This genotype variability reflected particular differences of Istrian cheese production, mainly variable salt concentration, ripening temperature and air humidity as well as microclimatic or vegetation conditions. There was considerable variation between the strains of the same species regarding wide range of biotechnologically important traits as well as their ability to survive in simulated gastrointestinal conditions. A considerable number of strains were resistant to critically important antibiotics such as tetracycline (43.56 %), erythromycin (35.79 %) and vancomycin (23.48 %). Polymerase chain reaction-based detection did not identify any of the common genetic determinants for vancomycin and erythromycin resistance; for tetracycline tetM gene was detected. The presence of virulence genes including agg, efaAfs, gelE, cylM, cylB, cylA, esp, efaAfm, cob and cpd was frequently recorded, especially among E. faecalis strains. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the safety and probiotic potential characteristics of ten Lactobacillus spp. strains ( Lactobacillus fermentum SJRP30, Lactobacillus casei SJRP37, SJRP66, SJRP141, SJRP145, SJRP146, and SJRP169, and Lactobacillus delbrueckii subsp. bulgaricus SJRP50, SJRP76, and SJRP149) that had previously been isolated from water buffalo mozzarella cheese. The safety of the strains was analyzed based on mucin degradation, hemolytic activity, resistance to antibiotics and the presence of genes encoding virulence factors. The in vitro tests concerning probiotic potential included survival under simulated gastrointestinal (GI) tract conditions, intestinal epithelial cell adhesion, the presence of genes encoding adhesion, aggregation and colonization factors, antimicrobial activity, and the production of the β-galactosidase enzyme. Although all strains presented resistance to several antibiotics, the resistance was limited to antibiotics to which the strains had intrinsic resistance. Furthermore, the strains presented a limited spread of genes encoding virulence factors and resistance to antibiotics, and none of the strains presented hemolytic or mucin degradation activity. The L. delbrueckii subsp. bulgaricus strains showed the lowest survival rate after exposure to simulated GI tract conditions, whereas all of the L. casei and L. fermentum strains showed good survivability. None of the tested lactobacilli strains presented bile salt hydrolase (BSH) activity, and only L. casei SJRP145 did not produce the β-galactosidase enzyme. The strains showed varied levels of adhesion to Caco-2 cells. None of the cell-free supernatants inhibited the growth of pathogenic target microorganisms. Overall, L. fermentum SJRP30 and L. casei SJRP145 and SJRP146 were revealed to be safe and to possess similar or superior probiotic characteristics compared to the reference strain L. rhamnosus GG (ATCC 53103). [ABSTRACT FROM AUTHOR]

The seasonal variation and removal efficiency of antibiotic resistance genes (ARGs), including tetracycline resistance genes ( tetG, tetM, and tetX) and macrolide ( ermB, ermF, ereA, and mefA), were investigated in two typical swine wastewater treatment systems in both winter and summer. ARGs, class 1 integron gene, and 16S rRNA gene were quantified using real-time polymerase chain reaction assays. There was a 0.31-3.52 log variation in ARGs in raw swine wastewater, and the abundance of ARGs in winter was higher than in summer. tetM, tetX, ermB, ermF, and mefA were highly abundant. The abundance of ARGs was effectively reduced by most individual treatment process and the removal efficiencies of ARGs were higher in winter than in summer. However, when examining relative abundance, the fate of ARGs was quite variable. Anaerobic digestion reduced the relative abundance of tetX, ermB, ermF, and mefA, while lagoon treatment decreased tetM, ermB, ermF, and mefA. Sequencing batch reactor (SBR) decreased tetM, ermB, and ermF, but biofilters and wetlands did not display consistent removal efficiency on ARGs in two sampling seasons. As far as the entire treatment system is concerned, ermB and mefA were effectively reduced in both winter and summer in both total and relative abundance. The relative abundances of tetG and ereA were significantly correlated with intI1 ( p < 0.01), and both tetG and ereA increased after wastewater treatment. This may pose a great threat to public health. [ABSTRACT FROM AUTHOR]

This study investigated the occurrence, antimicrobial resistance and virulence of Enterococcus from poultry and cattle farms. Three hundred and ninety samples: cloacal/rectal swabs ( n = 260) and manure ( n = 130] were processed for recovery of Enterococcus species. Standard bacteriological methods were used to isolate, identify and characterize Enterococcus species for antimicrobial susceptibility and expression of virulence traits. Detection of antibiotic resistance and virulence genes was carried out by polymerase chain reaction. Enterococcus was recovered from 167 (42.8%) of the 390 samples tested with a predominance of Enterococcus faecium (27.7%). Other species detected were Enterococcus gallinarum, Enterococcus faecalis, Enterococcus hirae, Enterococcus raffinosus, Enterococcus avium, Enterococcus casseliflavus, Enterococcus mundtii and Enterococcus durans. All the isolates tested were susceptible to vancomycin, but resistance to tetracycline, erythromycin, ampicillin and gentamicin was also observed among 61.0, 61.0, 45.1 and 32.7% of the isolates, respectively. Sixty (53.1%) of the isolates were multidrug resistant presenting as 24 different resistance patterns with resistance to gentamicin-erythromycin-streptomycin-tetracycline (CN-ERY-STR-TET) being the most common ( n = 11) pattern. In addition to expression of virulence traits (haemolysin, gelatinase, biofilm production), antibiotic resistance ( tetK, tetL, tetM, tetO and ermB) and virulence ( asa1, gelE, cylA) genes were detected among the isolates. Also, in vitro transfer of resistance determinants was observed among 75% of the isolates tested. Our data revealed poultry, cattle and manure in this area are hosts to varying Enterococcus species harbouring virulence and resistance determinants that can be transferred to other organisms and also are important for causing nosocomial infection. [ABSTRACT FROM AUTHOR]

The Epidemiology of otitis media with spontaneous perforation of the tympanic membrane and associated nasopharyngeal carriage of bacterial otopathogens was analysed in a county in Catalonia (Spain) with pneumococcal conjugate vaccines (PCVs) not included in the immunization programme at study time. A prospective, multicentre study was performed in 10 primary care centres and 2 hospitals (June 2011-June 2014), including all otherwise healthy children ≥2 months ≤8 years with otitis media presenting spontaneous tympanic perforation within 48h. Up to 521 otitis episodes in 487 children were included, showing by culture/PCR in middle ear fluid (MEF): Haemophilus influenzae [24.2%], both Streptococcus pneumoniae and H. influenzae [24.0%], S. pneumoniae [15.9%], Streptococcus pyogenes [13.6%], and Staphylococcus aureus [6.7%]. Culture-negative/PCR-positive otitis accounted for 31.3% (S. pneumoniae), 30.2% (H. influenzae) and 89.6% (mixed S. pneumoniae/H. influenzae infections). Overall, incidence decreased over the 3-year study period, with significant decreases in otitis by S. pneumoniae and by H. influenzae, but no decreases for mixed S. pneumoniae/H. influenzae infections. Concordance between species in nasopharynx and MEF was found in 58.3% of cases, with maximal rates for S. pyogenes (71.8%), and with identical pneumococcal serotype in 40.5% of cases. Most patients (66.6%) had past episodes. PCV13 serotypes were significantly more frequent in first episodes, in otitis by S. pneumoniae as single agent, and among MEF than nasopharyngeal isolates. All non-PCV13 serotypes separately accounted for <5% in MEF. Up to 73.9% children had received ≥1 dose of PCV, with lower carriage of PCV13 serotypes than among non-vaccinated children. Pooling pneumococcal isolates from MEF and nasopharynx, 30% were multidrug resistant, primarily belonging to serotypes 19A [29.8%], 24A [14.3%], 19F [8.3%] and 15A [6.0%]. Our results suggest that increasing PCV13 vaccination would further reduce transmission of PCV13 serotypes with special benefits for youngest children (with none or uncompleted vaccine schedules), preventing first otitis episodes and subsequent recurrences. [ABSTRACT FROM AUTHOR]