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Применение в хирургической практике антисептиков в значительной степени дополняет профилактику и лечение гнойно-воспалительных инфекций. Проблема широкого распространения устойчивых к антимикробным препаратам микроорганизмов осложняет борьбу с возбудителями инфекций, связанными с оказанием медицинской помощи. Необходимо детальное изучение бактериологической эффективности существующих препаратов для последующего дифференцированного применения. Цель исследования определить антимикробную активность современных антисептиков в отношении актуальных в хирургической практике полирезистентных госпитальных штаммов микроорганизмов и дрожжеподобного гриба C. albicans. Материалы и методы исследования. Методом серийных разведений в питательной среде, содержащей микроорганизмы, проведено исследование антимикробной активности современных антисептиков (диметилсульфоксид, коллоидное серебро, поли(1 -этенил-2-пирролидинон) йодид, бензилдиметил[3-(миристоиламино)пропил]аммонийхлорид, N, У-бис (4-хлорфенил) -3,12 -диимино -2,4,11,13 -тетраазатетрадекандиимидамид, полигексанид, гидроксиметилхи-ноксалиндиоксид) в отношении наиболее значимых возбудителей инфекции, связанной с оказанием медицинской помощи (K. pneumoniae, A. baumannii, P. aeruginosa, E. coli, S. aureus) и дрожжеподобного гриба C. albicans. Результаты и их обсуждение. Выявлены отличия в антисептическом эффекте применяемых в хирургическом стационаре препаратов. Максимальную универсальную антимикробную активность против грамотрицательных бактерий проявил гидроксиметилхиноксалиндиоксид, зона задержки роста бактерий составила 23-26 мм, а N, У-бис (4-хлорфенил)-3,12-диимино-2,4,11,13-тетраазатетрадекандиимидамид (11-19 мм) и полигексанид (14-16 мм) проявили более слабый противобактериальный эффект. Против грамположительного S. aureus все антисептики показали сопоставимую антибактериальную активность. Из всех изученных препаратов только полигексанид, хлоргексидин и бензилдиметил[3-(миристоиламино)пропил]аммонийхлорид действовали на дрожжеподобные грибы C. albicans. Заключение. Оценка бактериологической эффективности антисептических препаратов, применяемых в хирургии, показала разницу в степени выраженности их действия на возбудителей инфекций. Полученные данные по чувствительности возбудителей гнойно-воспалительных инфекций к современным антисептическим препаратам послужат повышению эффективности лечения послеоперационных ран., The use of antiseptics in surgical practice largely completes the prevention and treatment of purulent-inflammatory infections. The problem of widespread distribution of antimicrobial-resistant microorganisms complicates the fight against infectious agents associated with the provision of medical care. It is necessary to study in detail the bacteriological efficacy of existing drugs for subsequent differentiated use. Goal. To determine the antimicrobial activity of modern antiseptics against polyresistant hospital strains of microorganisms and yeast-like fungus C. albicans, which are relevant in surgical practice. Materials and methods of research. The antimicrobial activity of modern antiseptics (dimethyl sulfoxide, colloidal silver, poly(1-ethenyl-2-pyrrolidinone) iodide, benzyldimethyl[3-(myristoylamino) was studied by the method of serial dilutions in a nutrient medium containing microorganismspropyl]ammonium chloride, N, N"-bis (4-chlorophenyl)-3,12-diimino-2,4,11,13-tetraazatetradecandiimidamide, polyhexanide, hydroxymethylquinoxalindioxide) in relation to the most significant pathogens of infection associated with medical care (K. pneumoniae, A. baumannii, P. aeruginosa, E. coli, S. aureus) and yeast like the fungus C. albicans. Results. Differences in the antiseptic effect of the drugs used in the surgical hospital were revealed. hydroxymethylquinoxalindioxide showed the maximum universal antimicrobial activity against gram-negative bacteria, the bacterial growth delay zone was 23-26 mm, and N,N"-bis (4-chlorophenyl)-3.12-diimino-2,4,11,13-tetraazatetradecandiimidamide (11-19 mm) and polyhexanide (14-16 mm) showed a weaker antibacterial effect. Against gram-positive S. aureus, all antiseptics showed comparable antibacterial activity. Of all the studied drugs, only polyhexanide, chlorhexidine and benzyldimethyl[3-(myristoylamino)propyl]ammonium chloride acted on yeast-like fungi C. albicans. Conclusion. Evaluation of the bacteriological effectiveness of antiseptic drugs used in surgery showed a difference in the severity of their effect on infectious agents. The obtained data on the sensitivity of pathogens of purulent-inflammatory infections to modern antiseptic drugs will serve to increase the effectiveness of treatment of postoperative wounds.

Occurrence of multidrug resistant Enterobacteriaceae in livestock is of concern as they can spread to humans. A potential introduction route for these bacteria to livestock could be animal feed. We therefore wanted to identify if Escherichia spp., Enterobacter spp., Klebsiella spp., or Raoutella spp. with transferable resistance to extended spectrum cephalosporins, carbapenems or colistin could be detected in the environment at feed mills in Sweden. A second aim was to compare detected isolates to previous described isolates from humans and animals in Sweden to establish relatedness which could indicate a potential transmission between sectors and feed mills as a source for antibiotic resistant bacteria. However, no isolates with transferable resistance to extended-cephalosporins or colistin could be identified, but one isolate belonging to the Enterobacter cloacae complex was shown to be carbapenem-resistant and showing carbapenemase-activity. Based on sequencing by both short-read Illumina and long-read Oxford Nanopore MinIon technologies it was shown that this isolate was an E. asburiae carrying a blaIMI-2 gene on a 216 Kbp plasmid, designated pSB89A/IMI-2, and contained the plasmid replicons IncFII, IncFIB, and a third replicon showing highest similarity to the IncFII(Yp). In addition, the plasmid contained genes for various functions such as plasmid segregation and stability, plasmid transfer and arsenical transport, but no additional antibiotic resistance genes. This isolate and the pSB89A/IMI-2 was compared to three human clinical isolates positive for blaIMI-2 available from the Swedish antibiotic monitoring program Swedres. It was shown that one of the human isolates carried a plasmid similar with regards to gene content to the pSB89A/IMI-2 except for the plasmid transfer system, but that the order of genes was different. The pSB89A/IMI-2 did however share the same transfer system as the blaIMI-2 carrying plasmids from the other two human isolates. The pSB89A/IMI-2 was also compared to previously published plasmids carrying blaIMI-2, but no identical plasmids could be identified. However, most shared part of the plasmid transfer system and DNA replication genes, and the blaIMI-2 gene was located next the transcription regulator imiR. The IS3-family insertion element downstream of imiR in the pSB89A was also related to the IS elements in other blaIMI-carrying plasmids.

Women with cystic fibrosis (CF) have a significantly lower life expectancy compared to men, which is indicated by an earlier impairment of lung function due to chronic colonization with biofilm formed by Pseudomonas aeruginosa. There is growing evidence that blood serum concentrations of the steroid sex hormone estradiol (E2) correlate with the occurrence of pulmonary exacerbations in CF but also play a role in the mucoid switch of P. aeruginosa. This study aims to shed light on possible microbiological reasons for sexual dimorphism in CF by investigating the influence of E2 on biofilm formation of P. aeruginosa CF isolates. For this purpose, 10 CF isolates of the respiratory tract derived from different CF patients have been treated with E2 in a microtiter plate biofilm model. Biofilms have been examined by crystal violet assays, field emission scanning electron microscopy (FE-SEM), 3D laser scanning microscopy (LSM), and quorum sensing (QS) reporter assays of the supernatants taken from biofilms. This allowed us to simultaneously investigate the effects of E2 on attached biofilm mass, biofilm ultrastructure, and QS activity. Upon E2 treatment, six out of 10 investigated CF isolates showed an increase of attached biofilm mass, whereas biofilms from two tested non-CF laboratory strains (PAO1 and ATCC19660) did not. Moreover, FE-SEM and 3D LSM analyses of the E2 responsive CF biofilms revealed ultrastructural remodeling of biofilm structure at different scales with increased formation of prominent biofilm spots, enhanced coverage with extracellular polymeric substance (EPS), and extended average surface roughness. QS activity measurements performed in biofilm supernatants via luminescence acyl homoserine lactone (AHL) reporter assays further showed that E2 treatment may also modulate QS signaling, as shown in an E2 sensitive CF isolate. Together, our results suggest the biofilm modulating effects of E2 on various clinical CF isolates that are documented by both biomass and ultrastructural changes of biofilms. The gained new insight into the influence of steroid hormones on P. aeruginosa biofilm phenotypes might pave the way for novel future approaches in personalized medicine based on the patients’ sex and hormonal status.

Azole antifungals, including fluconazole, have long been the first-line antifungal agents in the fight against fungal infections. The emergence of drug-resistant strains and the associated increase in mortality from systemic mycoses has prompted the development of new agents based on azoles. We reported a synthesis of novel monoterpene-containing azoles with high antifungal activity and low cytotoxicity. These hybrids demonstrated broad-spectrum activity against all tested fungal strains, with excellent minimum inhibitory concentration (MIC) values against both fluconazole-susceptible and fluconazole-resistant strains of Candida spp. Compounds 10a and 10c with cuminyl and pinenyl fragments demonstrated up to 100 times lower MICs than fluconazole against clinical isolates. The results indicated that the monoterpene-containing azoles had much lower MICs against fluconazole-resistant clinical isolates of Candida parapsilosis than their phenyl-containing counterpart. In addition, the compounds did not exhibit cytotoxicity at active concentrations in the MTT assay, indicating potential for further development as antifungal agents.

Background: Lawsonia inermis (L. inermis) is perennial plant commonly called henna. It is frequently cultivated in Sudan. Beside its uses cosmetics for staining hands and as hairs dyes‚ it was reported to be useful in jaundice, enlargement of spleen, calculus affliction and skin disease. Method: This descriptive study was done during the period from December 2014 to April 2015 in order to determine the invitro antimicrobial activity of L. inermis (henna) leaves extract against standard and clinical isolates from wound swabs, including Staphylococcus aureus, Escherichia coli, Proteus species, Klebsiella pneumonia, and Pseudomonas aeruginosa. These organisms were collected from different hospitals in Khartoum State including: Soba University Hospital, Military Teaching Hospital, and Laboratory Management Center. The in vitro antimicrobial susceptibly testing was performed using cup plate diffusion method. The activity of L. inermis Linn leaves extract was controlled with four reference antibiotics including gentamicin, oxacillin, ciprofloxacin, and impinim. Results: When aqueous extract of L. inermis Linn examined against standard bacteria and clinical isolates result showed that all standard bacteria were inhibited at 100%, 50%, and 25% concentration. All clinical isolates were successfully inhibited at 100%, 50%, 25%, and 12.5%. In contrary, the activity of methanolic extract of L. inermis Linn against standard bacteria showed that all standard bacteria were inhibited at 100%, 50% concentration, However, the clinical isolates showed an inhibition rate various depending on the concentration of methanolic extract of L. inermis Linn with S. aureus being most sensitive isolate. Conclusion: We conclude that aqueous and methanolic extract of henna exhibited antimicrobial activity against all types of tested organisms both clinical and standard isolates. But the aqueous extract shows superior inhibition ability than the methanolic.

The spread of antibiotic-resistant Acinetobacter baumannii poses a significant threat to public health worldwide. This nosocomial bacterial pathogen can be associated with life-threatening infections, particularly in intensive care units. A. baumannii is mainly described as an extracellular pathogen with restricted survival within cells. This study shows that a subset of A. baumannii clinical isolates extensively multiply within nonphagocytic immortalized and primary cells without the induction of apoptosis and with bacterial clusters visible up to 48 h after infection. This phenotype was observed for the A. baumannii C4 strain associated with high mortality in a hospital outbreak and the A. baumannii ABC141 strain, which was isolated from the skin but was found to be hyperinvasive. Intracellular multiplication of these A. baumannii strains occurred within spacious single membrane-bound vacuoles, labeled with the lysosomal associate membrane protein (LAMP1). However, these compartments excluded lysotracker, an indicator of acidic pH, suggesting that A. baumannii can divert its trafficking away from the lysosomal degradative pathway. These compartments were also devoid of autophagy features. A high-content microscopy screen of 43 additional A. baumannii clinical isolates highlighted various phenotypes, and (i) the majority of isolates remained extracellular, (ii) a significant proportion was capable of invasion and limited persistence, and (iii) three more isolates efficiently multiplied within LAMP1-positive vacuoles, one of which was also hyperinvasive. These data identify an intracellular niche for specific A. baumannii clinical isolates that enables extensive multiplication in an environment protected from host immune responses and out of reach of many antibiotics. IMPORTANCE Multidrug-resistant Acinetobacter baumannii isolates are associated with significant morbidity and mortality in hospitals worldwide. Understanding their pathogenicity is critical for improving therapeutic management. Although A. baumannii can steadily adhere to surfaces and host cells, most bacteria remain extracellular. Recent studies have shown that a small proportion of bacteria can invade cells but present limited survival. We have found that some A. baumannii clinical isolates can establish a specialized intracellular niche that sustains extensive intracellular multiplication for a prolonged time without induction of cell death. We propose that this intracellular compartment allows A. baumannii to escape the cell’s normal degradative pathway, protecting bacteria from host immune responses and potentially hindering antibiotic accessibility. This may contribute to A. baumannii persistence, relapsing infections, and enhanced mortality in susceptible patients. A high-content microscopy-based screen confirmed that this pathogenicity trait is present in other clinical A. baumannii isolates. There is an urgent need for new antibiotics or alternative antimicrobial approaches, particularly to combat carbapenem-resistant A. baumannii. The discovery of an intracellular niche for this pathogen, as well as hyperinvasive isolates, may help guide the development of antimicrobial therapies and diagnostics in the future.

Legionnaires’ Disease (LD) is a severe pneumonia mainly caused in Europe by Legionella pneumophila serogroup 1 (Lp1). Sequence-based typing methods reveal that some sequence types (ST) are overrepresented in clinical samples such as ST1 and ST47, suggesting that some strains are more fit for infection than others. In the present study, a collection of 108 Lp1 clinical isolates were used to evaluate the strain-dependent immune responses from human macrophages. Clinical Lp1 isolates induced differential TNFα secretion from macrophages. ST1 isolates induced a significantly higher TNF-α secretion than non-ST1, whereas ST47 isolates induced a significantly lower TNF-α secretion than non-ST47 isolates. ST1 isolates induced a significantly higher cell death than ST47 isolates evaluated by lactate dehydrogenase activity (cytotoxicity) and caspase-3 activity (apoptosis). Treatment of macrophages with anti-TNF-α antibodies significantly reduced the cell death in macrophages infected with ST1 or ST47 strains. The TNF-α secretion was neither explained by a differential bacterial replication nor by the number or type (bystander or infected) of TNF-α producing cells following infection but by a differential response from macrophages. The Paris ST1 reference strain elicited a significantly higher TNF-α gene transcription and a higher induction of NF-κB signaling pathway than the Lorraine ST47 reference strain. Clinical Lp1 isolates induce a diverse immune response and cell death, which could be related to the genotype. The two predominant sequence-types ST1 and ST47 trigger opposite inflammatory response that could be related to the host susceptibility.

В первый год пандемии COVID-19 от инфекций, устойчивых к антимикробным препаратам погибли 29 400 человек, в 40% случаев это была внутрибольничная инфекция. Большинство исследований по проблеме сепсиса посвящены преимущественно частным вопросам диагностики и лечения либо проведены на специфических группах пациентов. В то время как работ по микробиологии возбудителей сепсиса немного, а они требуют регулярного обновления. Цель исследования изучить микробиологические особенности сепсиса в современной многопрофильной медицинской организации. Материалы и методы исследования. Результаты бактериологического обследования 93 пациентов с сепсисом, проходивших лечение в крупной многопрофильной больнице Санкт-Петербурга в 2021 году. Результаты и их обсуждение. В обследованном стационаре преобладали легочные (33,3%) и урологические (23,7%) формы сепсиса. Внебольничные формы заболевания встречались значительно чаще (73,1%). При всех формах сепсиса наблюдалась высокая частота встречаемости полирезистентных штаммов K. pneumonia (51,5%). Это приводит к необходимости выбора антибиотиков последнего поколения в виде препаратов для эмпирической антибактериальной терапии, назначаемой до получения результатов лабораторного анализа на резистентность микроорганизмов. Так, наиболее часто назначаемыми антибактериальными препаратами были: меропенем (26%), цефтриаксон (26%) и амикацин (23,6%). Кроме того, при анализе гемокультур, была отмечена высокая доля полирезистентных A. baumannii (43,7%) в общем числе грамотрицательных микроорганизмов, выделенных у пациентов с сепсисом. Заключение. Увеличивается доля полирезистентных штаммов грамотрицательных микроорганизмов у пациентов с сепсисом, это осложняет эмпирическою антибиотикотерапию. В этой связи, еще больше актуализируется роль своевременных и качественных бактериологических исследований, как для пациентов, так и для тщательного и систематического отслеживания видоизменяющегося «микробиологического пейзажа» медицинской организации., In the first year of the COVID-19 pandemic, 29400 people died from antimicrobial-resistant infections, 40% of which were nosocomial infections. Most studies on the problem of sepsis are devoted mainly to particular issues of diagnosis and treatment, or were carried out on specific groups of patients. While there are few works on the microbiology of sepsis pathogens, they require regular updating. The research purpose is to study the microbiological features of sepsis in a modern multidisciplinary medical organization. Materials and research methods. Results of a bacteriological examination of 93 patients with sepsis treated in a large multidisciplinary hospital in St. Petersburg in 2021. Results and its discussion. In the examined hospital, pulmonary (33.3%) and urological (23.7%) forms of sepsis prevailed. Community-acquired forms of the disease were much more common (73.1%). In all forms of sepsis, there was a high incidence of multiresistant strains of K. pneumonia (51.5%). This leads to the need to choose the latest generation of antibiotics in the form of drugs for empirical antibiotic therapy, prescribed before the results of laboratory analysis for microbial resistance. Thus, the most commonly prescribed antibacterial drugs were: meropenem (26%), ceftriaxone (26%) and amikacin (23.6%). In addition, when analyzing blood cultures, a high proportion of multi-resistant A. baumannii (43.7%) was noted in the total number of gram-negative microorganisms isolated from patients with sepsis. Conclusion. The proportion of multidrug-resistant strains of gram-negative microorganisms in patients with sepsis is increasing, which complicates empirical antibiotic therapy. In this regard, the role of timely and high-quality bacteriological studies, both for patients and for careful and systematic monitoring of the changing "microbiological landscape" of a medical organization, is even more relevant.

Бактериемия является одним из возможных признаков сепсиса в связи с чем, настоящее время возрастает актуальность бактериологического исследования образцов крови пациентов. Цель исследования. Дать характеристику видового разнообразия бактерий, выделенных из крови пациентов, определить спектр антибиотикорезистентности и охарактеризовать динамику выявленных изменений. Материалы и методы исследования. Проведен анализ результатов бактериологического исследования образцов крови пациентов. Результаты и их обсуждение. Дана сравнительная характеристика спектра бактерий, выделенных из крови пациентов многопрофильного стационара в 2016 - 2020 гг. Большинство изолятов выделено от пациентов хирургического профиля. Соотношение выделенных грамположительных и грамотрицательных бактерий изменялось за исследуемый период наблюдения. В 2017 году 59,1% бактериемии был обусловлен грамотрицательной флорой, в 2018 году 35,3%, в период пандемии в 2020 г. - 46,1%. Среди гемокультур преобладали штаммы коагулазонегативных стафилококков (28,5%) и Klebsiella pneumoniae (22,2%). Высокой оказалась также доля Enterococcus spp. (7,9%) и Acinetobacter baumannii (7,5%). Выводы. Ведущие возбудители инфекции кровотока среди грамотрицательных бактерий отличались высокой частотой полирезистентности к антибиотикам, включая карбапенемы. Отмечены возросшая роль K. pneumoniae и неферментирующих грамотрицательных бактерий в этиологии инфекций кровотока, а также высокие уровни их резистентности к антибактериальным средствам., Bacteremia is one of the possible signs of sepsis, and therefore, the relevance of bacteriological examination of patients' blood samples is currently increasing. The research purpose is to characterize the species diversity of bacteria isolated from the blood of patients, to determine the spectrum of antibiotic resistance and to characterize the dynamics of the identified changes. Materials and methods of research. The analysis of the results of bacteriological examination of patients' blood samples was carried out. Results and its discussion. A comparative characteristic of the spectrum of bacteria isolated from the blood of patients of a multidisciplinary hospital in 2016 - 2020 is given. Most of the isolates were isolated from surgical patients. The ratio of isolated gram - positive and gram - negative bacteria changed during the study period of observation. In 2017, 59.1% of bacteremia was caused by gram - negative flora, in 2018 - 35.3%, during the pandemic in 2020 - 46.1%. Among the hemocultures, strains of coagulase - negative staphylococci (28.5%) and Klebsiellapneumoniae (22.2%) prevailed. The proportion of Enterococcusspp (7.9%) and Acinetobacterbaumannii (7.5%) was also high. Conclusion. The leading pathogens of bloodstream infection among gram - negative bacteria were characterized by a high frequency of polyresistance to antibiotics, including carbapenems. The increased role of K. pneumoniae and non - fermenting gram - negative bacteria in the etiology of bloodstream infections, as well as high levels of their resistance to antibacterial agents, were noted.

Klebsiella pneumoniae is an opportunistic pathogen that frequently causes nosocomial and community-acquired (CA) infections. Until now, a limited number of studies has been focused on the analyses of changes affecting the virulence attributes. Genotypic and phenotypic methods were used to characterise the 39 clinical K. pneumoniae isolates; all belonged to the pan-drug resistant, widespread clone ST 15 and expressed the K24 capsule. PFGE has revealed that the isolates could be divided into three distinct genomic clusters. All isolates possessed allS and uge genes, known to contribute to the virulence of K. pneumoniae and 10.25% of the isolates showed hypermucoviscosity, 94.87% produced type 1 fimbriae, 92.3% produced type 3 fimbriae, and 92.3% were able to produce biofilm. In vivo persistence could be supported by serum resistance 46.15%, enterobactin (94.87%) and aerobactin (5.12%) production and invasion of the INT407 and T24 cell lines. Sequence analysis of the whole genomes of the four representative strains 11/3, 50/1, 53/2 and 53/3 has revealed high sequence homology to the reference K. pneumoniae strain HS11286. Our results represent the divergence of virulence attributes among the isolates derived from a common ancestor clone ST 15, in an evolutionary process that occurred both in the hospital and in the community.

Enterococcus faecalis and E. faecium are the major pathogens causing community- and healthcare-associated infections, with an ability to acquire resistance to multiple antimicrobials. The present study was conducted to determine the prevalence of virulence factors, drug resistance and its genetic determinants, and clonal lineages of E. faecalis and E. faecium clinical isolates in northern Japan. A total of 480 (426 E. faecalis and 54 E. faecium) isolates collected over a four-month period were analyzed. Three virulence factors promoting bacterial colonization (asa1, efaA, and ace) were more prevalent among E. faecalis (46–59%) than E. faecium, while a similar prevalence of enterococcal surface protein gene (esp) was found in these species. Between E. faecalis and E. faecium, an evident difference was noted for resistance to erythromycin, gentamicin, and levofloxacin and its responsible resistance determinants. Oxazolidinone resistance gene optrA and phenicol exporter gene fexA were identified in an isolate of E. faecalis belonging to ST480 and revealed to be located on a cluster similar to those of isolates reported in other Asian countries. The E. faecalis isolates analyzed were differentiated into 12 STs, among which ST179 and ST16 of clonal complex (CC) 16 were the major lineage. Nearly all the E. faecium isolates were assigned into CC17, which consisted of 10 different sequence types (STs), including a dominant ST17 containing multidrug resistant isolates and ST78 with isolates harboring the hyaluronidase gene (hyl). The present study revealed the genetic profiles of E. faecalis and E. faecium clinical isolates, with the first identification of optrA in ST480 E. faecalis in Japan.

Antimicrobial resistance (AMR) data from humans are mostly collected from clinical isolates, whereas from livestock data also exist from colonizing pathogens. In Germany, livestock data are collected from clinical and nonclinical isolates. We compared resistance levels of clinical and nonclinical isolates of Escherichia coli from weaning and fattening pigs with clinical outpatient isolates of humans from urban and rural areas. We also studied the association of AMR with available antimicrobial use (AMU) data from humans and pigs. Differences between rural and urban isolates were minor and did not affect the comparison between human and pig isolates. We found higher resistance levels to most antimicrobials in human isolates compared to nonclinical isolates of fattening pigs. Resistance to ampicillin, however, was significantly more frequent in clinical isolates of fattening pigs and in clinical and nonclinical isolates of weaning pigs compared to isolates from humans. The opposite was observed for ciprofloxacin. Co-trimoxazole resistance proportions were higher in clinical isolates of weaning and fattening pigs as compared to isolates from humans. Resistance proportions were higher in clinical isolates than in nonclinical isolates from pigs of the same age group and were also higher in weaner than in fattening pigs. Significant associations of AMU and AMR were found for gentamicin resistance and aminoglycoside use in humans (borderline) and for ampicillin resistance in clinical isolates and penicillin use in fattening pigs. In summary, we found significant differences between isolates from all populations, requiring more detailed analyses supported by molecular data and better harmonized data on AMU and AMR.

Clinical management of human immunodeficiency virus type-1 (HIV-1) infection may be negatively impacted by either acquired or transmitted drug resistance. Here, we aim to extend our understanding of the impact of resistance-associated mutations (RAMs) on the susceptibility of clinical isolates to the nonnucleoside reverse transcriptase inhibitor (NNRTI) doravirine. Clinical isolates from people living with HIV-1 undergoing routine testing for susceptibility to doravirine and other approved NNRTIs (etravirine, rilpivirine, efavirenz, and nevirapine) were collected from August 2018 to August 2019. Susceptibility in the presence/absence of NNRTI and nucleos(t)ide reverse transcriptase inhibitor (NRTI) mutations was determined using cutoffs for relative fold change in inhibition (ratio of the 50% inhibitory concentration [IC 50 ] of patient virus compared with the IC 50 of a wild-type reference strain). Biological cutoffs of 3- to 15-fold change were investigated for doravirine, with preestablished cutoffs used for the other NNRTIs. Of 4,070 clinical isolates, 42.9% had ≥1 NNRTI RAM. More isolates were susceptible to doravirine (92.5–96.7%) than to etravirine (91.5%), rilpivirine (89.5%), efavirenz (81.5%), or nevirapine (77.5%). Based on a 3-fold cutoff, doravirine susceptibility was retained in 44.7–65.8% of isolates resistant to another NNRTI and 28.5% of isolates resistant to all other tested NNRTIs. The presence of NRTI RAMs including thymidine analog mutations was associated with doravirine hypersusceptibility in some isolates, particularly in the absence of NNRTI RAMs. These results support the favorable resistance profile of doravirine and are of particular importance given the challenge posed by both acquired and transmitted resistance.

Among non-tuberculous mycobacteria, Mycobacterium kansasii is one of the most pathogenic, able to cause pulmonary disease indistinguishable from tuberculosis in immunocompetent susceptible adults. The lack of animal models that reproduce human-like lung disease, associated with the necrotic lung pathology, impairs studies of M. kansasii virulence and pathogenicity. In this study, we examined the ability of the C57BL/6 mice, intratracheally infected with highly virulent M. kansasii strains, to produce a chronic infection and necrotic lung pathology. As a first approach, we evaluated ten M. kansasii strains isolated from Brazilian patients with pulmonary disease and the reference strain M. kansasii ATCC 12478 for virulence-associated features in macrophages infected in vitro; five of these strains differing in virulence were selected for in vivo analysis. Highly virulent isolates induced progressive lung disease in mice, forming large encapsulated caseous granulomas in later stages (120–150 days post-infection), while the low-virulent strain was cleared from the lungs by day 40. Two strains demonstrated increased virulence, causing premature death in the infected animals. These data demonstrate that C57BL/6 mice are an excellent candidate to investigate the virulence of M. kansasii isolates. We observed considerable heterogeneity in the virulence profile of these strains, in which the presence of highly virulent strains allowed us to establish a clinically relevant animal model. Comparing public genomic data between Brazilian isolates and isolates from other geographic regions worldwide demonstrated that at least some of the highly pathogenic strains isolated in Brazil display remarkable genomic similarities with the ATCC strain 12478 isolated in the United States 70 years ago (less than 100 SNPs of difference), as well as with some recent European clinical isolates. These data suggest that few pathogenic clones have been widely spread within M. kansasii population around the world.

Listeria monocytogenes is a foodborne human pathogen that affects public health worldwide. Whole-genome sequencing (WGS) can classify L. monocytogenes isolates and identify virulence islands and resistance genes potentially influencing infectivity. Herein, WGS was used to assess 151 L. monocytogenes isolates from 120 cases of clinical infection in Beijing, China, between 2014 and 2018. Most isolates were either serogroup 1/2a,3a or serogroup 1/2b,3b,7, with 25 multilocus sequence typing (MLST) types (STs) represented, of which ST8, ST87, and ST5 were the most common. Core-genome MLST (cgMLST) grouped the 151 isolates into 116 cgMLST types. The discriminatory power of cgMLST was greater than other subtypes, revealing that isolates from the same patient were highly related (only differing at one allele). Eighty-six isolates formed 30 complexes with ≤ 7 cgMLST alleles between neighboring isolates, suggesting possible outbreaks. Compared with isolates in the United States, ST8, ST121, ST619, ST87, and ST155 isolates were grouped into unified clades. All 151 isolates were positive for common virulence-associated loci, and 26 lineage I isolates harbored the pathogenicity island 3 (LIPI-3) locus, while 42 lineage I isolates harbored the complete LIPI-4 locus. Eleven ST619 isolates had both LIPI-3 and LIPI-4. Among the 151 isolates, 13 were resistant to at least one antibiotic, and no multidrug-resistant isolates were identified. Resistance phenotypes correlated with genotypes, apart from two meropenem resistance isolates. The findings provided insight into the nature of L. monocytogenes strains currently causing clinical disease in Beijing, and WGS analysis indicated possible outbreaks.

Globally, tuberculosis (TB) remains a prevalent threat to public health. In 2019, TB affected 10 million people and caused 1.4 million deaths. The major challenge for controlling this infectious disease is the emergence and spread of drug-resistant Mycobacterium tuberculosis, the causative agent of TB. The antibiotic streptomycin is not a current first-line anti-TB drug. However, WHO recommends its use in patients infected with a streptomycin-sensitive strain. Several mutations in the M. tuberculosis rpsL, rrs and gidB genes have proved association with streptomycin resistance. In this study, we performed a molecular analysis of these genes in clinical isolates to determine the prevalence of known or novel mutations. Here, we describe the genetic analysis outcome. Furthermore, a biocomputational analysis of the MtGidB L101F variant, the product of a novel mutation detected in gidB during molecular analysis, is also reported as a theoretical approach to study the apparent genotype-phenotype association.

To survive when confronted by antibiotics, some strains of microorganisms have evolved and developed the ability to resist the most commonly prescribed forms of medication. As a result, the search for plant derived substances are now becoming one of major areas of research. Plants and their parts are valuable for food, shelter, nutraceutical and major constituent of drugs used in traditional herbal medicinal systems.Caladium bicolorleaf is used in African traditional medicine for the treatment of infections in Nigeria. In this study, the active compounds ofCaladium bicolorleaf were extracted using methanol and investigated for the presence of secondary metabolites. Antimicrobial activity of the extract was determined against selected clinical isolates (Staphylococcus aureus,Escherichia coli,Proteus mirabilis,Klebsiella pneumoniae,Aspergillus fumigatusandCandida albicans) using the agar well diffusion. The methanol extract demonstrated a significant broad-spectrum antimicrobial activity against both gram-positive and gram-negative bacteria, with the highest activity having a zone of inhibition of 10 mm. The antimicrobial activities could be attributed to the presence of phytochemicals compounds such as tannins, alkaloids, saponins. This is interesting and promising and may therefore be exploited further to understand better the mechanisms responsible for the antimicrobial activity recorded. This study can also serve as scientific bases for support of the traditional use ofCaladium bicolorleaf in treatment of infections.

Among non-tuberculous mycobacteria, Mycobacterium kansasii is one of the most pathogenic, able to cause pulmonary disease indistinguishable from tuberculosis in immunocompetent susceptible adults. The lack of animal models that reproduce human-like lung disease, associated with the necrotic lung pathology, impairs studies of M. kansasii virulence and pathogenicity. In this study, we examined the ability of the C57BL/6 mice, intratracheally infected with highly virulent M. kansasii strains, to produce a chronic infection and necrotic lung pathology. As a first approach, we evaluated ten M. kansasii strains isolated from Brazilian patients with pulmonary disease and the reference strain M. kansasii ATCC 12478 for virulence-associated features in macrophages infected in vitro; five of these strains differing in virulence were selected for in vivo analysis. Highly virulent isolates induced progressive lung disease in mice, forming large encapsulated caseous granulomas in later stages (120–150 days post-infection), while the low-virulent strain was cleared from the lungs by day 40. Two strains demonstrated increased virulence, causing premature death in the infected animals. These data demonstrate that C57BL/6 mice are an excellent candidate to investigate the virulence of M. kansasii isolates. We observed considerable heterogeneity in the virulence profile of these strains, in which the presence of highly virulent strains allowed us to establish a clinically relevant animal model. Comparing public genomic data between Brazilian isolates and isolates from other geographic regions worldwide demonstrated that at least some of the highly pathogenic strains isolated in Brazil display remarkable genomic similarities with the ATCC strain 12478 isolated in the United States 70 years ago (less than 100 SNPs of difference), as well as with some recent European clinical isolates. These data suggest that few pathogenic clones have been widely spread within M. kansasii population around the world.

Background The ability to culture Plasmodium falciparum continuously in vitro has enabled stable access to asexual and sexual parasites for malaria research. The portfolio of isolates has remained limited and research is still largely based on NF54 and its derived clone 3D7. Since 1978, isolates were collected and cryopreserved at Radboudumc from patients presenting at the hospital. Here, procedures are described for culture adaptation of asexual parasites, cloning and production of sexual stage parasites responsible for transmission (gametocytes) and production of oocysts in Anopheles mosquitoes. This study aimed to identify new culture-adapted transmissible P. falciparum isolates, originating from distinct geographical locations. Methods Out of a collection of 121 P. falciparum isolates stored in liquid nitrogen, 21 from different geographical origin were selected for initial testing. Isolates were evaluated for their ability to be asexually cultured in vitro, their gametocyte production capacity, and consistent generation of oocysts. Results Out of 21 isolates tested, twelve were excluded from further analysis due to lack of mature gametocyte production (n = 1) or generation of satisfactory numbers of oocysts in mosquitoes (n = 11). Nine isolates fulfilled selection criteria and were cloned by limiting dilution and retested. After cloning, one isolate was excluded for not showing transmission. The remaining eight isolates transmitted to Anopheles stephensi or Anopheles coluzzii mosquitoes and were categorized into two groups with a reproducible mean oocyst infection intensity above (n = 5) or below five (n = 3). Conclusions These new P. falciparum culture-adapted isolates with reproducible transmission to Anopheles mosquitoes are a valuable addition to the malaria research tool box. They can aid in the development of malaria interventions and will be particularly useful for those studying malaria transmission.

Vibrio parahaemolyticus is a common foodborne pathogen that causes gastroenteritis worldwide. Determining its prevalence and genetic diversity will minimize the risk of infection and the associated economic burden. Multilocus sequence typing (MLST) is an important tool for molecular epidemiology and population genetic studies of bacteria. Here, we analyzed the genetic and evolutionary relationships of 162 V. parahaemolyticus strains isolated in the Guangdong Province, China, using MLST. In the study, 120 strains were isolated from food samples, and 42 strains were isolated from clinical samples. All strains were categorized into 100 sequence types (STs), of which 58 were novel (48 from the food isolates and 10 from the clinical isolates). ST415 was the most prevalent ST among the food isolates, while ST3 was the most prevalent ST among the clinical isolates. Further, 12 clonal complexes, 14 doublets, and 73 singletons were identified in all ST clusters, indicating high genetic diversity of the analyzed strains. At the concatenated sequence level, non-synonymous sites in both, food and clinical isolates, were associated with purifying selection. Of note, the dN/dS ration was greater than 1 for some housekeeping genes in all isolates. This is the first time that some loci under positive selection were identified. These observations confirm frequent recombination events in V. parahaemolyticus. Recombination was much more important than mutation for genetic heterogeneity of the food isolates, but the probabilities of recombination and mutations were almost equal for the clinical isolates. Based on the phylogenetic analysis, the clinical isolates were concentrated in the maximum-likelihood tree, while the food isolates were heterogeneously distributed. In conclusion, the food and clinical isolates of V. parahaemolyticus from the Guangdong Province are similar, but show different evolutionary trends. This may help prevent large-scale spread of highly virulent strains and provides a genetic basis for the discovery of microevolutionary relationships in V. parahaemolyticus populations.

International audience; Livestock data on antimicrobial resistance (AMR) are commonly collected from bacterial populations of clinical and non-clinical isolates. In contrast to data on non-clinical isolates from livestock, data on clinical isolates are not harmonized in Europe. The Normalized Resistance Interpretation (NRI) method was applied to overcome the lack of harmonization of laboratory methods and interpretation rules between monitoring systems. Statistical analyses were performed to identify associations between the isolate type (clinical vs. non-clinical) and resistance to four antimicrobials (ampicillin, tetracycline, gentamicin, and nalidixic acid) per animal category in Germany and France. Additional statistical analyses comparing clinical and non-clinical isolates were performed with the available data on the same antimicrobial panel and animal categories from the UK and Norway. Higher resistance prevalence was found in clinical isolates compared to non-clinical isolates from calves to all antimicrobials included in Germany and France. It was also found for gentamicin in broilers from France. In contrast, in broilers and turkeys from Germany and France and in broilers from the UK, a higher resistance level to ampicillin and tetracycline in non-clinical isolates was encountered. This was also found in resistance to gentamicin in isolates from turkeys in Germany. Resistance differed within countries and across years, which was partially in line with differences in antimicrobial use patterns. Differences in AMR between clinical and non-clinical isolates of Escherichia coli are associated with animal category (broiler, calf, and turkey) and specific antimicrobials. The NRI method allowed comparing results of non-harmonized AMR systems and might be useful until international harmonization is achieved.

Urogenital Chlamydia trachomatis infection is the most common sexually transmitted bacterial infection throughout the world. While progress has been made to better understand how type strains develop and respond to environmental stress in vitro, very few studies have examined how clinical isolates behave under similar conditions. Here, we examined the development and persistence phenotypes of several clinical isolates, to determine how similar they are to each other, and the type strain C. trachomatis D/UW-3/Cx. The type strain was shown to produce infectious progeny at a higher magnitude than each of the clinical isolates, in each of the six tested cell lines. All chlamydial strains produced the highest number of infectious progeny at 44 h post-infection in the McCoy B murine fibroblast cell line, yet showed higher levels of infectivity in the MCF-7 human epithelial cell line. The clinical isolates were shown to be more susceptible than the type strain to the effects of penicillin and iron deprivation persistence models in the MCF-7 cell line. While subtle differences between clinical isolates were observed throughout the experiments conducted, no significant differences were identified. This study reinforces the importance of examining clinical isolates when trying to relate in vitro data to clinical outcomes, as well as the importance of considering the adaptations many type strains have to being cultured in vitro.

In India, multidrug resistance determinants are much more abundant in community-associated bacterial pathogens due to the improper treatment of domestic and industrial effluents. In particular, a high bacterial load of the opportunistic pathogen P. aeruginosa in sewage and water bodies in India is well documented., Multidrug-resistant community-acquired infections caused by the opportunistic human pathogen Pseudomonas aeruginosa are increasingly reported in India and other locations globally. Since this organism is ubiquitous in the environment, samples such as sewage and wastewater are rich reservoirs of P. aeruginosa bacteriophages. In this study, we report the isolation and characterization of a novel P. aeruginosa N4-like lytic bacteriophage, vB_Pae_AM.P2 (AM.P2), from wastewater in Kerala, India. AM.P2 is a double-stranded DNA podovirus that efficiently lyses the model strain, PAO1, at a multiplicity of infection as low as 0.1 phage per bacterium and resistance frequency of 6.59 × 10−4. Synergy in bactericidal activity was observed between AM.P2 and subinhibitory concentrations of the antibiotic ciprofloxacin. Genome sequencing of AM.P2 revealed features similar to those of the N4-like P. aeruginosa phages LUZ7 and KPP21. As judged by two independent assay methods, spot tests and growth inhibition, AM.P2 successfully inhibited the growth of almost 30% of strains from a contemporary collection of multidrug-resistant P. aeruginosa clinical isolates from South India. Thus, AM.P2 may represent an intriguing candidate for inclusion in bacteriophage cocktails developed for various applications, including water decontamination and clinical bacteriophage therapy. IMPORTANCE In India, multidrug resistance determinants are much more abundant in community-associated bacterial pathogens due to the improper treatment of domestic and industrial effluents. In particular, a high bacterial load of the opportunistic pathogen P. aeruginosa in sewage and water bodies in India is well documented. The isolation and characterization of bacteriophages that could target emerging P. aeruginosa strains, representing possible epicenters for community-acquired infections, could serve as a useful alternative tool for various applications, such as phage therapy and environmental treatment. Continuing to supplement the repertoire of broad-spectrum bacteriophages is an essential tool in confronting this problem.

Polymorphonuclear leukocytes (PMNs) are regarded as vehicles for the hematogenous dissemination of human cytomegalovirus (HCMV). In cell culture, this concept has been validated with cell-free laboratory strains but not yet with clinical HCMV isolates that grow strictly cell-associated. We, therefore, aimed to evaluate whether PMNs can also transmit such isolates from initially infected fibroblasts to other cell types, which might further clarify the role of PMNs in HCMV dissemination and provide a model to search for potential inhibitors. PMNs, which have been isolated from HCMV-seronegative individuals, were added for 3 h to fibroblasts infected with recent cell-associated HCMV isolates, then removed and transferred to various recipient cell cultures. The transfer efficiency in the recipient cultures was evaluated by immunofluorescence staining of viral immediate early antigens. Soluble derivatives of the cellular HCMV entry receptor PDGFRα were analyzed for their potential to interfere with this transfer. All of five tested HCMV isolates could be transferred to fibroblasts, endothelial and epithelial cells with transfer rates ranging from 2 to 9%, and the transferred viruses could spread focally in these recipient cells within 1 week. The PDGFRα-derived peptides IK40 and GT40 reduced transfer by 40 and 70% when added during the uptake step. However, when added during the transfer step, only IK40 was effective, inhibiting transmission by 20% on endothelial cells and 50–60% on epithelial cells and fibroblasts. These findings further corroborate the assumption of cell-associated HCMV dissemination by PMNs and demonstrate that it is possible to inhibit this transmission mode.

The study aimed to investigate the potency of Leptospermum petersonii, Eucalyptus citriodora, Pelargonium graveolens, and Pelargonium roseum essential oils (EOs) against Streptococcus sanguinis and Streptococcus salivarius dental isolates. To examine the variation in sensitivity among strains of different origin, isolates from symptomatic and asymptomatic root canal infections, as well as the reference, strains S. sanguinis ATCC 10556 and S. salivarius ATCC 9222 were used in the study. EOs were chemically characterised (GC/MS). Their antibacterial and antibiofilm properties were detected by microdilution and CV assay respectively. Statistical analysis was performed by One way ANOVA. Examined bacterial strains, regardless clinical or ATCC, exhibited the highest sensitivity to L. petersonii and E. citriodora EOs (MIC ranged 0.53±0.00 - 5.88±1.27 mg/ml and 0.08±0.03 - 9.37±1.25 mg/ml respectively), while two other EOs were less efficient. Comparison of MIC values showed that sensitivity of the isolates recovered from symptomatic infections was the least, while the susceptibility of asymptomatic isolates and especially of reference strains were higher. EO inhibition of biofilm formation differed between clinical isolates of S. salivarius and their reference strain. Inhibition was higher in the case of S. salivarius ATCC 9222 (84 %-89 %) in comparison to that of clinical isolates (24 %-75 %). Regardless of the strains tested, E. citriodora exhibited the highest antibiofilm potential. To conclude: L. petersonii induced notable antibacterial potential, while E. citriodora EO proved promising antibacterial and antibiofilm activity. Due to the variable sensitivity of the isolates to EOs, it is suggested to screen activity on a large number of strains, especially from symptomatic infections.

Green synthesis of nanoparticles is a widely researched and popular direction in the development of nanotechnology. It is a simple, cheap and effective method for obtaining nanoparticles with interesting biological properties. In light of the development of antibiotic resistance to important clinical strains of bacteria, this method was used in the present study to obtain silver nanoparticles with antibacterial activity. The aim of this study was to synthesise silver nanoparticles with antibacterial action by yeast in a process known as “green synthesis”. We are also considering the prospect of using silver nanoparticles as an antibacterial substance for drug development. The production of nanoparticles was confirmed by UV spectroscopy. Staphylococcus aureus ATCC 25923 and Escherichia coli ATCC 25922 test strains and Staphylococcus aureus 1536 and Klebsiella pneumoniae 520 clinical isolates were used to study the antibacterial effect. The effect of synthesised nanoparticles on the metabolic activity of bacterial cells and their ability to adhere, as well as the minimum inhibitory concentrations (MICs) of synthesised nanoparticles for each of the strains, were determined. Following UV spectroscopy, the nanoparticles obtained were found to have a pronounced peak in optical absorption at 400 nm, corresponding to the plasmon resonance of silver nanoparticles, and demonstrated a high antibacterial effect against all the strains studied.

Non-typhoidal Salmonella present a major threat to animal and human health as food-borne infectious agents. We characterized 91 bacterial isolates from Armenia and Georgia in detail, using a suite of assays including conventional microbiological methods, determining antimicrobial susceptibility profiles, matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry, serotyping (using the White-Kauffmann-Le Minor scheme) and genotyping (repetitive element sequence-based PCR (rep-PCR)). No less than 61.5% of the isolates were shown to be multidrug-resistant. A new antimicrobial treatment strategy is urgently needed. Phage therapy, the therapeutic use of (bacterio-) phages, the bacterial viruses, to treat bacterial infections, is increasingly put forward as an additional tool for combatting antibiotic resistant infections. Therefore, we used this representative set of well-characterized Salmonella isolates to analyze the therapeutic potential of eleven single phages and selected phage cocktails from the bacteriophage collection of the Eliava Institute (Georgia). All isolates were shown to be susceptible to at least one of the tested phage clones or their combinations. In addition, genome sequencing of these phages revealed them as members of existing phage genera (Felixounavirus, Seunavirus, Viunavirus and Tequintavirus) and did not show genome-based counter indications towards their applicability against non-typhoidal Salmonella in a phage therapy or in an agro-food setting.

Salmonella is a foodborne pathogen affecting over 200 million people and resulting in over 200,000 fatal cases per year. Its adhesion to and invasion into intestinal epithelial cells represent one of the first and key steps in the pathogenesis of salmonellosis. Still, around 35 to 40% of bacterial genes have no experimentally validated function, and their contribution to bacterial virulence, including adhesion and invasion, remains largely unknown. Therefore, the significance of this study is in the identification of new genes or gene allelic variants previously not associated with adhesion and invasion. It is well established that blocking adhesion and/or invasion would stop or hamper bacterial infection; therefore, the new findings from this study could be used in future developments of anti-Salmonella therapy targeting genes involved in these key processes. Such treatment could be a valuable alternative, as the prevalence of antibiotic-resistant bacteria is increasing very rapidly., The initial steps of Salmonella pathogenesis involve adhesion to and invasion into host epithelial cells. While well-studied for Salmonella enterica serovar Typhimurium, the factors contributing to this process in other, host-adapted serovars remains unexplored. Here, we screened clinical isolates of serovars Gallinarum, Dublin, Choleraesuis, Typhimurium, and Enteritidis for adhesion to and invasion into intestinal epithelial cell lines of human, porcine, and chicken origins. Thirty isolates with altered infectivity were used for genomic analyses, and 14 genes and novel mutations associated with high or low infectivity were identified. The functions of candidate genes included virulence gene expression regulation and cell wall or membrane synthesis and components. The role of several of these genes in Salmonella adhesion to and invasion into cells has not previously been investigated. The genes dksA (encoding a stringent response regulator) and sanA (encoding a vancomycin high-temperature exclusion protein) were selected for further analyses, and we confirmed their roles in adhesion to and invasion into host cells. Furthermore, transcriptomic analyses were performed for S. Enteritidis and S. Typhimurium, with two highly infective and two marginally infective isolates for each serovar. Expression profiles for the isolates with altered infection phenotypes revealed the importance of type 3 secretion system expression levels in the determination of an isolate’s infection phenotype. Taken together, these data indicate a new role in cell host infection for genes or gene variants previously not associated with adhesion to and invasion into the epithelial cells. IMPORTANCE Salmonella is a foodborne pathogen affecting over 200 million people and resulting in over 200,000 fatal cases per year. Its adhesion to and invasion into intestinal epithelial cells represent one of the first and key steps in the pathogenesis of salmonellosis. Still, around 35 to 40% of bacterial genes have no experimentally validated function, and their contribution to bacterial virulence, including adhesion and invasion, remains largely unknown. Therefore, the significance of this study is in the identification of new genes or gene allelic variants previously not associated with adhesion and invasion. It is well established that blocking adhesion and/or invasion would stop or hamper bacterial infection; therefore, the new findings from this study could be used in future developments of anti-Salmonella therapy targeting genes involved in these key processes. Such treatment could be a valuable alternative, as the prevalence of antibiotic-resistant bacteria is increasing very rapidly.

To investigate the prevalence, probable mechanisms and serotype correlation of colistin resistance in clinical isolates of Salmonella from patients in China, Salmonella isolates were collected from fecal and blood samples of patients. In this study, 42.8% (136/318) clinical isolated Salmonella were resistant to colistin. MIC distribution for colistin at serotype level among the two most prevalent serotypes originating from humans in China indicated that Salmonella Enteritidis (83.9% resistance, 125/149) were significantly less susceptible than Salmonella Typhimurium (15.3% resistance, 9/59, P < 0.01). mcr genes and mutations in PmrAB confer little for rate of colistin resistant Salmonella isolated from human patients. Phylogenetic tree based on core-genome single nucleotide polymorphisms (SNPs) was separately by the serotypes and implied a diffused distribution of MICs in the same serotype isolates. Relatvie expression levels of colistin resistant related pmr genes were significantly higher in non-mcr colistin resistant S. Typhimurium than in colistin sensitive S. Typhimurium, but no discernable differences between colistin resistant and sensitive S. Enteritidis, indicating a different mechanism between colistin resistant S. Typhimurium and S. Enteritidis. In conclusion, colistin susceptibility and colistin resistant mechanism of clinical isolated Salmonella were closely associated with specific serotypes, at least in the two most prevalent serotype Enteritidis and Typhimurium. We suggest clinical microbiology laboratory interpreting Salmonella colistin MIC results in the serotype level.

Mycobacterium tuberculosis (Mtb) infects alveolar macrophages (AMs), causing pulmonary tuberculosis (PTB), the most common form of the disease. Less frequently, Mtb is disseminated to many other organs and tissues, resulting in different extrapulmonary forms of TB. Nevertheless, very few studies have addressed the global mRNA response of human AMs, particularly from humans with the active form of the disease. Strikingly, almost no studies have addressed the response of human extrapulmonary macrophages to Mtb infection. In this pilot study, using microarray technology, we examined the transcriptomic ex vivo response of AMs from PTB patients (AMTBs) and AMs from control subjects (AMCTs) infected with two clinical isolates of Mtb. Furthermore, we also studied the infection response of human splenic macrophages (SMs) to Mtb isolates, as a model for extrapulmonary infection, and compared the transcriptomic response between AMs and SMs. Our results showed a striking difference in global mRNA profiles in response to infection between AMs and SMs, implicating a tissue-specific macrophage response to Mtb.

There are still major gaps in our understanding of the bacterial factors that influence the outcomes of human Campylobacter jejuni infection. The aim of this study was to compare the virulence-associated features of 192 human C. jejuni strains isolated from hospitalized patients with diarrhoea (150/192, 78.1%), bloody diarrhoea (23/192, 11.9%), gastroenteritis (3/192, 1.6%), ulcerative colitis (3/192, 1.5%), and stomach ache (2/192, 1.0%). Traits were analysed with genotypic and phenotypic methods, including PCR and extracellular matrix protein (ECMP) binding, adhesion, and invasion capacities. Results were studied alongside patient symptoms, but no distinct links with them could be determined. Since the capacity of C. jejuni to invade host epithelial cells is one of its most enigmatic attributes, a high throughput transcriptomic analysis was performed in the third hour of internalization with a C. jejuni strain originally isolated from bloody diarrhoea. Characteristic groups of genes were significantly upregulated, outlining a survival strategy of internalized C. jejuni comprising genes related (1) to oxidative stress; (2) to a protective sheath formed by the capsule, LOS, N-, and O- glycosylation systems; (3) to dynamic metabolic activity supported by different translocases and the membrane-integrated component of the flagellar apparatus; and (4) to hitherto unknown genes.

Close to half of a collection of 27 clinical a/α isolates of Candida albicans underwent white-to-opaque switching. Complementation experiments revealed that while approximately half of the a/α switchers were due to EFG1 mutations, the remaining half were due to mutations in other genes. In addition, the results of competition experiments in a mouse GI tract colonization model support previous observations that efg1/efg1 cells rapidly outcompete EFG1/EFG1 strains, but direct microscopic analysis reveals that the major colonizing cells were opaque, not gray., The transcription factor EFG1 functions as a suppressor of white-to-opaque and white-to-gray switching in a/α strains of Candida albicans. In a collection of 27 clinical isolates, 4 of the 17 EFG1/EFG1 strains, 1 of the 2 EFG1/efg1 strains, and all 8 of the efg1/efg1 strains underwent white-to-opaque switching. The four EFG1/EFG1 strains, the one EFG1/efg1 strain, and one of the eight efg1/efg1 strains that underwent switching to opaque did not switch to gray and could not be complemented with a copy of EFG1. Competition experiments in a mouse model for gastrointestinal (GI) colonization confirmed that efg1/efg1 cells rapidly outcompete EFG1/EFG1 cells, and in plating experiments, formed colonies containing both gray and opaque cells. Direct microscopic analysis of live cells in the feces, however, revealed that the great majority of cells were opaque, suggesting opaque, not gray, may be the dominant phenotype at the site of colonization. IMPORTANCE Close to half of a collection of 27 clinical a/α isolates of Candida albicans underwent white-to-opaque switching. Complementation experiments revealed that while approximately half of the a/α switchers were due to EFG1 mutations, the remaining half were due to mutations in other genes. In addition, the results of competition experiments in a mouse GI tract colonization model support previous observations that efg1/efg1 cells rapidly outcompete EFG1/EFG1 strains, but direct microscopic analysis reveals that the major colonizing cells were opaque, not gray.

Limited therapy options due to antibiotic resistance underscore the need for optimization of current diagnostics. In some bacterial species, antimicrobial resistance can be unambiguously predicted based on their genome sequence. In this study, we sequenced the genomes and transcriptomes of 414 drug‐resistant clinical Pseudomonas aeruginosa isolates. By training machine learning classifiers on information about the presence or absence of genes, their sequence variation, and expression profiles, we generated predictive models and identified biomarkers of resistance to four commonly administered antimicrobial drugs. Using these data types alone or in combination resulted in high (0.8–0.9) or very high (> 0.9) sensitivity and predictive values. For all drugs except for ciprofloxacin, gene expression information improved diagnostic performance. Our results pave the way for the development of a molecular resistance profiling tool that reliably predicts antimicrobial susceptibility based on genomic and transcriptomic markers. The implementation of a molecular susceptibility test system in routine microbiology diagnostics holds promise to provide earlier and more detailed information on antibiotic resistance profiles of bacterial pathogens and thus could change how physicians treat bacterial infections., The spread of antibiotic resistance complicates infection treatment, requiring an optimization of current diagnostics. In this study, a machine learning approach identified a set of biomarkers suitable for the development of a molecular test system to determine antibiotic resistance profiles.

Intracellular invasion is an advantageous mechanism used by pathogens to evade host defense and antimicrobial therapy. In patients, the intracellular microbial lifestyle can lead to infection persistence and recurrence, thus worsening outcomes. Lung infections caused by Pseudomonas aeruginosa, especially in cystic fibrosis (CF) patients, are often aggravated by intracellular invasion and persistence of the pathogen. Proliferation of the infectious species relies on a continuous deoxyribonucleotide (dNTP) supply, for which the ribonucleotide reductase enzyme (RNR) is the unique provider. The large genome plasticity of P. aeruginosa and its ability to rapidly adapt to different environments are challenges for studying the pathophysiology associated with this type of infection. Using different reference strains and clinical isolates of P. aeruginosa independently combined with alveolar (A549) and bronchial (16HBE14o- and CF-CFBE41o-) epithelial cells, we analyzed host–pathogen interactions and intracellular bacterial persistence with the aim of determining a cell type-directed infection promoted by the P. aeruginosa strains. The oscillations in cellular toxicity and oxygen consumption promoted by the intracellular persistence of the strains were also analyzed among the different infectious lung models. Significantly, we identified class II RNR as the enzyme that supplies dNTPs to intracellular P. aeruginosa. This discovery could contribute to the development of RNR-targeted strategies against the chronicity occurring in this type of lung infection. Overall our study demonstrates that the choice of bacterial strain is critical to properly study the type of infectious process with relevant translational outcomes.

Drug resistance of Pseudomonas aeruginosa is a leading problem in hospital infections. The aim of this study was to determine the best molecular genetic discrimination method for Pseudomonas spp. isolates among 94 outpatients and inpatients and see their grouping by phenotype characteristics (biofilm formation, frequency of serotypes, pigmentation, production of different class of beta-lactamases, and susceptibility to different antibiotic classes) and genotype. The most common serotypes were P1, P6, and P11, while co-productions of pyoverdine and pyocyanin were observed in 70 % of isolates. A total of 77.66 % isolates were mostly weak and moderate biofilm producers. Isolates were susceptible to colistin (100 %), aztreonam (97.87 %), imipenem (91.49 %), doripenem (90.43 %), and meropenem (84.04 %). MICs values confirmed susceptibility to ceftazidime and cefepime and singled out meripenem as the most effective inhibitor. Most isolates were resistant to aminoglycosides and fluoroquinolones. Only two isolates produced ESBL, eight were carbapenemase producers, and five isolates produced MBLs. Twenty-nine isolates were multidrug-resistant; 82.8 % of which produced both pigments, 58.3 % were non-typeable, while the P6 and P11 serotypes were equally distributed (16.7 %). Thirteen MDR isolates were strong enzyme producers. RAPD PCR analysis using primer 272 proved the best at discriminatory fingerprinting for Pseudomonas isolates, as it allocated 12 clusters. A correlation between DNA patterns and antibiotic resistance, production of pigments, serotypes distribution, and biofilm formation was not observed, and only confirmed higher genetic heterogeneity among P. aeruginosa isolates, which suggests that other molecular methods are needed to reveal potential relations between genotypic patterns and phenotypic characteristics., Antibiotska rezistencija Pseudomonas aeruginosa vodeći je problem u bolničkim infekcijama. Cilj ovoga istraživanja bio je utvrditi najbolju diskriminatorno molekularno-genetičku metodu među 94 ambulantna i bolnička Pseudomonas spp. izolata kako bi se uvidjelo njihovo grupiranje u smislu različitih fenotipskih obilježja (stvaranje biofilma, učestalost serotipova, pigmentacija, proizvodnja različitih klasa beta-laktamaza i osjetljivost na različite skupine antibiotika) u skladu s genotipom. Najčešći serotipovi bili su P1, P6 i P11, a proizvodnja i pioverdina i piocijanina primijećena je kod 70 % izolata. Ukupno 77,66 % izolata uglavnom je iskazalo slabu i umjerenu proizvodnju biofilma. Izolati su bili osjetljivi na kolistin (100 %), aztreonam (97,87 %), imipenem (91,49 %), doripenem (90,43 %) i meropenem (84,04 %). Vrijednosti MIC-ova potvrdile su podložnost izolata ceftazidimu i cefepimu, a izdvojile su meropenem kao najučinkovitiji inhibitor. Većina izolata bila je otporna na aminoglikozid i fluorokinolon. Samo dva izolata proizvela su ESBL, osam izolata sintetiziralo je karbapenemaze, a pet izolata imalo je sposobnost proizvodnje MBL-a. Dvadeset devet izolata bilo je višestruko rezistentno na antibiotike, od kojih je 82,8 % proizvodilo oba pigmenta, 58,3 % bili su netipabilni, a serotipovi P6 i P11 bili su podjednako zastupljeni među njima (16,7 %). Trinaest MDR izolata bili su snažni proizvođači enzima. RAPD PCR analiza korištenjem 272 početnica pokazala se kao najbolja diskriminatorna metoda otiskom prsta (fingerprinting) za Pseudomonas izolate, izdvajajući čak 12 različitih klastera. U ovom istraživanju nije zabilježena povezanost između DNA obrazaca i otpornosti na antibiotike, proizvodnje pigmenata, distribucije serotipova i stvaranja biofilma, što potvrđuje puno veću genetičku heterogenost unutar samih izolata P. aeruginosa, pod čim se podrazumijeva uključivanje drugih molekularnih metoda u otkrivanju potencijalnih odnosa između genetičkih obrazaca i fenotipskih obilježja.

Introduction: The clinical significance of enterococci is mostly related to its antibiotic resistance which contributes to colonization and infection, in particular amongst the hospitalized patients. The present review has examined the literature to provide a comprehensive data on enterococci antibiotic resistance during the last 20 years in Iran. Methods: Search engines such as Google Scholar and PubMed were used to identify all Persian and English-language articles investigating enterococci in Iran from 1996 to 2017. The search terms were “enterococci”, “enterococcal”, “enterococcus”, “Iran”, “bacterial resistance”, “antibiotic resistance” and “resistance”. Results: Decrease in the resistance trend against ampicillin, gentamycin and ciprofloxacin was observed over a period of 15 years (2001 to 2016) in Iran. During a 10 years period from 2001 to 2015, the rate of resistance among Enterococcus faecalis species was less than Enterococcus faecium. The resistancerate, however, was considerably increased for both species during this period. The mean resistance rates for vancomycin, gentamicin, ciprofloxacin, erythromycin, nitrofurantoin, chloramphenicol, trimethoprim-sulfametoxazol, imipenem and teicoplanin were higher among complicated cases (patients with underlying debilitating disorders) compared to general cases (hospitalized or outpatients with no specific underlying disorder). Conclusions: E. faecalis and E. faecium showed a rise in the mean resistance against all the antibiotics during a 10-year period from 2010 to 2015. With the exception of penicillin and ampicillin, resistance to all antibiotics was higher amongst complicated cases compared to general patients. Keywords: Antimicrobial resistance, clinical isolates, enterococci, Iran, review, resistance trend

The antibacterial activity of plant extracts of the Lamiaceae family was evaluated against clinical isolates of multi-resistant Gram-negative bacteria by broth microdilution technique. Promising results were obtained considering that all extracts were active for at least two bacterial species with MIC ranging from 0.5 to 2.0 mg/mL.

Malaria remains a major cause of childhood deaths in resource-limited settings. In the absence of an effective vaccine, drugs and other interventions have played very significant roles in combating the scourge of malaria. The recent reports of resistance to artemisinin necessitate the need for new antimalarial drugs with novel mechanisms of action. Towards the development of new, affordable and easily accessible antimalarial drugs for endemic regions, the Medicines for Malaria Venture (MMV) assembled a total of 400 active antimalarial compounds called the Malaria Box. The potency and the efficacy of the Malaria Box Compounds have been determined mainly using laboratory strains of P. falciparum. This study investigated the potency of twenty compounds from the Malaria Box against four clinical isolates from Ghana, using optimized in vitro growth inhibitory assays. Seven out of the 20 compounds screened had 50% inhibitory concentration (IC50) below 500 nM. The most active among the selected compounds was MMV006087 (average IC50 of 30.79 nM). Variations in the potency of the Malaria Box Compounds were observed between P. falciparum clinical isolates and Dd2 strain. We also investigated the sensitivity of the clinical isolates to chloroquine and artesunate. The N093 clinical isolate was found to be resistant to chloroquine but showed high sensitivity to artesunate. The results underscore the importance of including clinical isolates with different drug-resistant backgrounds, in addition to laboratory strains, in validating potential compounds during antimalarial compound screening programs., Graphical abstract Image 1, Highlights • MMV006087, MMV085203, MMV008956 and MMV665977 were most potent compounds in this study. • MMV006087 was the best compound in this study. • N093 clinical isolate harboured K76T mutation and was sensitive to artesunate but resistant to chloroquine. • Malaria Box Compounds have varied potency in clinical isolates and laboratory strains of P. falciparum.

Globally, tuberculosis (TB) remains a prevalent threat to public health. In 2019, TB affected 10 million people and caused 1.4 million deaths. The major challenge for controlling this infectious disease is the emergence and spread of drug-resistant Mycobacterium tuberculosis, the causative agent of TB. The antibiotic streptomycin is not a current first-line anti-TB drug. However, WHO recommends its use in patients infected with a streptomycin-sensitive strain. Several mutations in the M. tuberculosisrpsL, rrs and gidB genes have proved association with streptomycin resistance. In this study, we performed a molecular analysis of these genes in clinical isolates to determine the prevalence of known or novel mutations. Here, we describe the genetic analysis outcome. Furthermore, a biocomputational analysis of the MtGidB L101F variant, the product of a novel mutation detected in gidB during molecular analysis, is also reported as a theoretical approach to study the apparent genotype-phenotype association.

Background: Hospital-acquired infections are among the most significant reasons of human mortality world-wide which can be controlled by efficient application of suitable disinfectant for hospital setting. The main goal of the present study was to determine the efficacy of eight routinely used hospital disinfectants against clinical isolates. Methods: In our descriptive study, in the first step the antibiogram assay of 99 clinical isolates enterococci and Pseudomonas aeruginosa were determined. Then, minimum inhibitory concentration and minimum bactericidal concentration of isolates against Povidone Iodine 10%, Ethanol 70%, Savlon 3.2%, Deconex51Gastro, Procept Floor, Septo med, Surfanious and Gigasept AF were evaluated. Furthermore, the efficacy of disinfectants was reevaluated in presence of 5% (w/v) Bovine Serum Albumin. Results: The results showed that Septo med and Surfanious are the most and less potent disinfectants against clinical isolates, respectively. It is also resulted that Povidone Iodine is the most effective choice among the conventional disinfectants in this study. Clearly, addition of 5% organic substances reduced the efficacy of selected disinfectants significantly. Conclusion: Novel quaternary ammonium compounds are the most applicable choice for disinfection of hospital surfaces and instruments in this study.

Background. The spirochete Borrelia miyamotoi belongs to the group of borrelia causing tick-borne relapsing fevers. It is transmitted by the same tick species as Borrelia burgdorferi sensu lato and is widespread in temperate climate zones of Eurasia and North America. Previously we have shown by PCR that clinical cases presented as systemic febrile illness and diagnosed usually as «Ixodes tick-borne borreliosis without erythema migrans» were caused by B. miyamotoi. According to recent estimation in Russia there are several thousands human cases of B. miyamotoi infection annually. The investigation of this emerging human pathogen was hampered because the attempts to culture this bacterium directly from human blood failed. Results and conclusion: We here describe protocol - avoiding the inhibitory effects of anticoagulants on B. miyamotoi growth - enabling successful B. miyamotoi isolation from blood of Russian patients.

Item does not contain fulltext The spread of multidrug resistant Mycobacterium tuberculosis is one of the major challenges in tuberculosis control. In Eurasia, the spread of multidrug resistant tuberculosis is driven by the M. tuberculosis Beijing genotype. In this study, we examined whether selective advantages are present in the proteome of Beijing isolates that contribute to the emergence of this genotype. To this end, we compared the proteome of M. tuberculosis Beijing to that of M. tuberculosis H37Rv, both in the presence and absence of the first-line antibiotic rifampicin. During rifampicin exposure, both M. tuberculosis genotypes express proteins belonging to the DosR dormancy regulon, which induces a metabolically hypoactive-, drug tolerant phenotype. However, these markers of rifampicin tolerance were already more abundant in the M. tuberculosis Beijing isolate prior to drug exposure. To determine whether the a priori high abundance of specific proteins contribute to the formation of antibiotic resistance in M. tuberculosis Beijing, we quantified the abundance of 33 selected proteins in 27 clinical isolates from the five most common M. tuberculosis lineages using parallel reaction monitoring. The observed pre-existing high abundance of dormancy proteins in Beijing strains provides an evolutionary advantage that allows these strains to persist for prolonged periods during rifampicin treatment. SIGNIFICANCE: M. tuberculosis is the leading cause of death by a bacterial infection worldwide. Treatment-regimen to eradicate this pathogen make use of the first-line antibiotic rifampicin, which is considered to be the cornerstone of modern day anti-tuberculosis treatment. Despite the potency of rifampicin, there is an increasing occurrence of rifampicin resistant mutants in a specific cluster of M. tuberculosis, the Beijing genotype. Using both a data dependent acquisition and a targeted proteomic approach we identified markers of rifampicin tolerance to be high abundant in members of the M. tuberculosis Beijing genotype, already prior drug exposure. The identification of this M. tuberculosis Beijing specific trait will contribute to improved diagnostics and treatment of M. tuberculosis.

Monocytes play a critical role during infection with Mycobacterium tuberculosis (Mtb). They are recruited to the lung, where they participate in the control of infection during active tuberculosis (TB). Alternatively, inflammatory monocytes may participate in inflammation or serve as niches for Mtb infection. Monocytes response to infection may vary depending on the particularities of the clinical isolate of Mtb from which they are infected. In this pilot study, we have examined the baseline mRNA profiles of circulating human monocytes from patients with active TB (MoTB) compared with monocytes from healthy individuals (MoCT). Circulating MoTB displayed a pro-inflammatory transcriptome characterized by increased gene expression of genes associated with cytokines, monocytopoiesis, and down-regulation of MHC class II gene expression. In response to in vitro infection with two clinical isolates of the LAM family of Mtb (UT127 and UT205), MoTB displayed an attenuated inflammatory mRNA profile associated with down-regulation the TREM1 signaling pathway. Furthermore, the gene expression signature induced by Mtb UT205 clinical strain was characterized by the enrichment of genes in pathways and biological processes mainly associated with a signature of IFN-inducible genes and the inhibition of cell death mechanisms compared to MoTB-127, which could favor the establishment and survival of Mtb within the monocytes. These results suggest that circulating MoTB have an altered transcriptome that upon infection with Mtb may help to maintain chronic inflammation and infection. Moreover, this functional abnormality of monocytes may also depend on potential differences in virulence of circulating clinical strains of Mtb.

Streptococcus dysgalactiae subsp. equisimilis (SDSE) is an emerging human pathogen that causes severe invasive streptococcal diseases. Recent reports have shown that SDSE exhibits high pathogenicity with different mechanisms from that of Streptococcus pyogenes, although the two streptococci possess some common virulence factors such as streptolysin, streptokinase, and cell-binding proteins. To date, only a few studies have examined the variety of mechanisms expressing the pathogenicity of SDSE. Among nine SDSE clinical isolates sequenced in this study, we present in vitro and in vivo analyses of KNZ01 and KNZ03, whose emm and multilocus species types (MLSTs) are prevalent in Japan and other countries. For the comparison of pathogenicity, we also utilized the ATCC 12394 strain. The whole-genome analysis showed that KNZ03 and ATCC 12394 are categorized into an identical clonal complex by MLST and are phylogenetically close. However, the three strains exhibited different characteristics for pathogenicity in vitro; ATCC 12394 showed significant cytotoxicity to human keratinocytes and release of streptolysin O (SLO) compared to KNZ01 and KNZ03; KNZ03 exhibited significantly high hemolytic activity, but did not secrete SLO. KNZ01 and KNZ03 adhered to human keratinocytes at a higher rate than ATCC 12394; KNZ03 showed a higher rate of survival after a brief (30 min) incubation with human neutrophils compared to the other two strains; also, KNZ01 grew more rapidly in the presence of human serum. In vivo subcutaneous infection commonly resulted in ulcer formation in the three strains 7 days after infection. KNZ01-infected mice showed significant body weight loss 2 days after infection. Besides, on post-infection day 2, only KNZ01 remained in the cutaneous tissues of mice. Scanning electron microscopy analysis revealed that KNZ01 formed an extracellular structure (biofilm), which was probably composed of cell wall-anchoring proteins, in the presence of glucose and human serum. The extracellular structure of ATCC 12394 was also changed dramatically in response to culture conditions, whereas that of KNZ03 did not. Our study proposed that each SDSE strain possesses different virulence factors characteristics for mediating pathogenicity in humans.

Bacteria often exist in polymicrobial communities where they compete for limited resources. Intrinsic to this competition is the ability of some species to inhibit or kill their competitors. This phenomenon is pervasive throughout the human body where commensal bacteria block the colonization of incoming microorganisms. In this regard, molecular epidemiological and microbiota-based studies suggest that species-specific interactions play a critical role in the prevention of nasal colonization of the opportunistic pathogen Staphylococcus aureus. Despite this, S. aureus exists as part of the microbiota of ∼25% of the population, suggesting that the interplay between S. aureus and commensals can be complex. Microbiota studies indicate that several bacterial genera are negatively correlated with S. aureus colonization. While these studies paint a broad overview of bacterial presence, they often fail to identify individual species-specific interactions; a greater insight in this area could aid the development of novel antimicrobials. As a proof of concept study designed to identify individual bacterial species that possess anti-S. aureus activity, we screened a small collection of clinical isolates from the Walter Reed National Military Medical Center for the ability to inhibit multiple S. aureus strains. We found that the majority of the isolates (82%) inhibited at least one S. aureus strain; 23% inhibited all S. aureus strains tested. In total, seven isolates mediated inhibitory activity that was independent of physical contact with S. aureus, and seven isolates mediated bactericidal activity. 16S rRNA based-sequencing revealed that the inhibitory isolates belonged to the Acinetobacter, Agromyces, Corynebacterium, Microbacteria, Mycobacterium, and Staphylococcus genera. Unexpectedly, these included seven distinct Acinetobacter baumannii isolates, all of which showed heterogeneous degrees of anti-S. aureus activity. Defined mechanistic studies on specific isolates revealed that the inhibitory activity was retained in conditioned cell free medium (CCFM) derived from the isolates. Furthermore, CCFM obtained from S. saprophyticus significantly decreased mortality of S. aureus-infected Galleria mellonella caterpillars. While future studies will seek to define the molecular mechanisms of the inhibitory activities, our current findings support the study of polymicrobial interactions as a strategy to understand bacterial competition and to identify novel therapeutics against S. aureus and other pathogens.

Xue Li,1,* Qi Luo,1,* Xinyu Yu,1 Yanling Zhang,1 Xiaoyue Cao,1 Dan Li2,3 1Non-Coding RNA and Drug Discovery Key Laboratory of Sichuan Province, Chengdu Medical College, Chengdu, Sichuan 610500, People’s Republic of China; 2School of Medical Laboratory Science, Chengdu Medical College, Chengdu, Sichuan 610500, People’s Republic of China; 3Sichuan Provincial Engineering Laboratory for Prevention and Control Technology of Veterinary Drug Residue in Animal-Origin Food, Chengdu Medical College, Chengdu, Sichuan 610500, People’s Republic of China*These authors contributed equally to this workCorrespondence: Dan LiSchool of Medical Laboratory Science, Chengdu Medical College, Chengdu, Sichuan 610500, People’s Republic of ChinaTel/Fax +86 28 62739520Email lidan0811@cmc.edu.cnBackground: Escherichia coli strains are the most commonly isolated bacteria in hospitals. The normally harmless commensal E. coli can become a highly adapted pathogen, capable of causing various diseases both in healthy and immunocompromised individuals, by acquiring a combination of mobile genetic elements. Our aim was to characterize E. coli strains from a hospital in western China to determine their virulence and antimicrobial resistance potential.Methods: A total of 97 E. coli clinical isolates were collected from the First Affiliated Hospital of Chengdu Medical College from 2015 to 2016. Microbiological methods, PCR, and antimicrobial susceptibility tests were used in this study.Results: The frequency of occurrence of the virulence genes fimC, irp2, fimH, fyuA, lpfA, hlyA, sat, and cnf1 in the E. coli isolates was 93.81, 92.78, 91.75, 84.54, 41.24, 32.99, 28.86, and 7.22%, respectively. Ninety-five (97.9%) isolates carried two or more different virulence genes. Of these, 44 (45.4%) isolates simultaneously harbored five virulence genes, 24 (24.7%) isolates harbored four virulence genes, and 17 (17.5%) isolates harbored six virulence genes. In addition, all E. coli isolates were multidrug resistant and had a high degree of antimicrobial resistance.Conclusion: These results indicate a high frequency of occurrence and heterogeneity of virulence gene profiles among clinical multidrug resistant E. coli isolates. Therefore, appropriate surveillance and control measures are essential to prevent the further spread of these isolates in hospitals.Keywords: Escherichia coli, clinical isolates, virulence genes, antimicrobial resistance, MDR

Background Herbaspirillum seropedicae is an environmental β-proteobacterium that is capable of promoting the growth of economically relevant plants through biological nitrogen fixation and phytohormone production. However, strains of H. seropedicae have been isolated from immunocompromised patients and associated with human infections and deaths. In this work, we sequenced the genomes of two clinical strains of H. seropedicae, AU14040 and AU13965, and compared them with the genomes of strains described as having an environmental origin. Results Both genomes were closed, indicating a single circular chromosome; however, strain AU13965 also carried a plasmid of 42,977 bp, the first described in the genus Herbaspirillum. Genome comparison revealed that the clinical strains lost the gene sets related to biological nitrogen fixation (nif) and the type 3 secretion system (T3SS), which has been described to be essential for interactions with plants. Comparison of the pan-genomes of clinical and environmental strains revealed different sets of accessorial genes. However, antimicrobial resistance genes were found in the same proportion in all analyzed genomes. The clinical strains also acquired new genes and genomic islands that may be related to host interactions. Among the acquired islands was a cluster of genes related to lipopolysaccharide (LPS) biosynthesis. Although highly conserved in environmental strains, the LPS biosynthesis genes in the two clinical strains presented unique and non-orthologous genes within the genus Herbaspirillum. Furthermore, the AU14040 strain cluster contained the neuABC genes, which are responsible for sialic acid (Neu5Ac) biosynthesis, indicating that this bacterium could add it to its lipopolysaccharide. The Neu5Ac-linked LPS could increase the bacterial resilience in the host aiding in the evasion of the immune system. Conclusions Our findings suggest that the lifestyle transition from environment to opportunist led to the loss and acquisition of specific genes allowing adaptations to colonize and survive in new hosts. It is possible that these substitutions may be the starting point for interactions with new hosts. Electronic supplementary material The online version of this article (10.1186/s12864-019-5982-9) contains supplementary material, which is available to authorized users.

The heterogeneity of the clinical outcome of Mycobacterium tuberculosis (Mtb) infection may be due in part to different strategies used by circulating strains to cause disease. This heterogeneity is one of the main limitations to eradicate tuberculosis disease. In this study, we have compared the transcriptional response of two closely related Colombian clinical isolates (UT127 and UT205) of the LAM family under two axenic media conditions. These clinical isolates are phenotypically different at the level of cell death, cytokine production, growth kinetics upon in vitro infection of human tissue macrophages, and membrane vesicle secretion upon culture in synthetic medium. Using RNA-seq, we have identified different pathways that account for two different strategies to cope with the stressful condition of a carbon-poor media such as Sauton’s. We showed that the clinical isolate UT205 focus mainly in the activation of virulence systems such as the ESX-1, synthesis of diacyl-trehalose, polyacyl-trehalose, and sulfolipids, while UT127 concentrates its efforts mainly in the survival mode by the activation of the DNA replication, cell division, and lipid biosynthesis. This is an example of two Mtb isolates that belong to the same family and lineage, and even though they have a very similar genome, its transcriptional regulation showed important differences. This results in summary highlight the necessity to reach a better understanding of the heterogeneity in the behavior of these circulating Mtb strains which may help us to design better treatments and vaccines and to identify new targets for drugs.

Supplemental Digital Content is available in the text, Burkholderia pseudomallei is the causative agent of meliodosis, and the cases in China are gradually increasing. The present retrospective study aimed to surveil the molecular epidemiological characteristics and antibiotic resistance of B pseudomallei isolates. B pseudomallei strains were isolated and verified from meliodosis patients with relevant epidemiological information from 2004 to 2016 in Hainan, China. Pulsed-field gel electrophoresis based on Spe I digestion was carried out, and antimicrobial resistance of B pseudomallei strains was observed against 9 frequently-used antimicrobials. A total of 164 B pseudomallei isolates were successfully divided into 60 pulsed-field gel electrophoresis (PFGE) patterns, including 33 clusters and 27 single types, at an 85% similarity level. The isolates also exhibited a high level of ceftazidime resistance rate (12.8%, 21/164). B pseudomallei strains were mainly heterogenous with no predominant type, but there were some clonal populations, dominate clusters prevalent and the resistance rates of cephems antimicrobial increased significantly between 2004 and 2016 along with the number of melioidosis cases collected in Hainan (cefoperazone-sulbactam [SCF], rs = 0.96, P = .04; ceftazidime [CAZ], rs = 0.98, P = .01). In conclusion, this study will help to enhance our understanding of molecular characteristics and antibiotic resistance of B pseudomallei.

Gram-negative (GN) rods cause about 10% periprosthetic joint infection (PJI) and represent an increasing challenge due to emergence of antimicrobial resistance. Escherichia coli and Pseudomonas aeruginosa are among the most common cause of GN-PJI and ciprofloxacin is the first-line antibiotic. Due to emergence of fluoroquinolone resistance, we evaluated in vitro the activity of fosfomycin, ciprofloxacin, and gentamicin, alone and in combinations, against E. coli and P. aeruginosa biofilms. Conventional microbiological tests and isothermal microcalorimetry were applied to investigate the anti-biofilm activity of the selected antibiotics against standard laboratory strains as well as clinical strains isolated from patients with prosthetic joint associated infections. The biofilm susceptibility to each antibiotic varied widely among strains, while fosfomycin presented a poor anti-biofilm activity against P. aeruginosa. Synergism of two-pair antibiotic combinations was observed against different clinical strains from both species. Highest synergism was found for the fosfomycin/gentamicin combination against the biofilm of E. coli strains (75%), including a gentamicin-resistant but fosfomycin-susceptible strain, whereas the gentamicin/ciprofloxacin combination presented synergism with higher frequency against the biofilm of P. aeruginosa strains (71.4%). A hypothetical bacteriolysis effect of gentamicin could explain why combinations with this antibiotic seem to be particularly effective. Still, the underlying mechanism of the synergistic effect on biofilms is unknown. In conclusion, combinatorial antibiotic application has shown to be more effective against biofilms compared to monotherapy. Further in vivo and clinical studies are essential to define the potential treatment regimen based on our results.