ermB Gene Research Articles

Phenotypic and genotypic characterization of macrolide resistance in Staphylococcus aureus isolates from wound infection

Microbiologists are increasingly concerned about the rise in S. aureus MLSB (macrolide, lincosamide, streptogramin B) drug resistance. Clindamycin has been effective in treating infections by S. aureus, and the variations in clindamycin sensitivity patterns cause treatment to fail. Inducible clindamycin resistance in S. aureus is expressed via erythromycin ribosome methylase genes. In the current study, 25 S. aureus isolates were identified by conventional chemical tests and the Vitek®2 system. Specific primers were used for the amplification of Macrolide genes by PCR. Among 25 S. aureus isolates, 23(92%) isolates were methicillin resistant and 2(8%) isolates were methicilin sensitive. The 5(20%) isolates showed resistance to Erythromycin and sensitivity to Clindamycin with a positive D test which was identified as inducible MLSB, while 2(8%) isolates showed resistance criteria for both Erythromycin and Clindamycin which identity as a constitutive MLSB and 18(92%) isolates were given the sensitivity for both Erythromycin and Clindamycin. The erm resistance genes (ermA, ermB, ermC, ermF, and ermG) were detected in 5(20%), 17(68%), 25(100%), 24(96%),11(44%) respectively. The D-test, and Vitek ®2 system should be routinely done to avoid treatment failure due to clindamycin resistance.

Taxonomical Identification and Safety Characterization of Lactobacillaceae from Mediterranean Natural Fermented Sausages.

Fermented meat products represent an important industrial sector in Europe, particularly in the Mediterranean Countries (MC), where the presence of numerous local productions, still obtained through spontaneous fermentation, is recognized as a formidable treasure chest of unexplored microbial biodiversity. Lactobacillaceae naturally occurring in fifteen spontaneously fermented sausages from MC (Italy, Spain, Croatia, and Slovenia) were isolated and taxonomically characterized using molecular techniques. Additionally, a safety assessment for the presence of antibiotic resistances and biogenic amine (BA) production was performed to determine their suitability as autochthonous starter cultures. Molecular typing, performed using REP-PCR, discriminated 151 strains belonging to Latilactobacillus sakei (59.6%), Latilactobacillus curvatus (26.5%) and Companilactobacillus alimentarius (13.9%). The minimum inhibitory concentrations (MICs) of eight different antibiotics revealed a high resistance to streptomycin (27%), tetracycline (16%), followed by gentamycin (14%) and kanamycin (13%). Interestingly, the results showed a geographical distribution of resistant biotypes. tetM/tetS or ermB genes were identified in only six strains. The amino-biogenic potential of the strains was assessed, confirming the absence of this trait among L. sakei, while a high number of producer strains was found among L. curvatus. On the 151 analyzed strains, 45 demonstrated safety traits for their future use as starter food cultures. These results open the way to further studies on the technological properties of these promising autochthonous strains, strongly linked to the Mediterranean environment.

Antibiotic Resistance Profile of RT 027/176 Versus Other Clostridioides difficile Isolates in Silesia, Southern Poland.

Clostridioides difficile is an important health care-associated pathogen. The aim of this study was to analyze the antibiotic susceptibility of C. difficile isolates from feces of patients from 13 hospitals in Silesia, Poland. The incidence of CDI per 100.000 people in Silesia in 2018–2019 was higher than the average in Poland (39.3–38.7 vs. 30.2–29.5, respectively). The incidence doubled from 26.4 in 2020 to 55.1 in 2021. Two hundred and thirty stool samples tested positive for GDH (glutamate dehydrogenase) and toxins were cultured anaerobically for C. difficile. The isolates were characterized, typed, and tested for susceptibility to 11 antibiotics by E-test (EUCAST, 2021). The genes of toxins A/B and binary were detected by mPCR. Of 215 isolates, 166 (77.2%) were classified as RT 027 and 6 (2.8%) as related RT 176. Resistance to ciprofloxacin (96.7%), moxifloxacin (79.1%), imipenem (78.1%), penicillin (67%), and rifampicin (40.5%) was found. The ermB gene was detected in 79 (36.7%) strains. Multidrug resistance (MDR) was confirmed in 50 (23.3%) strains of RT 027 (94%). We concluded that a high prevalence of MDR among hypervirulent RT 027/176 C. difficile was found in the Silesian region of Poland, emphasizing the need to enhance regional infection control on CDI and antibiotic stewardships.

Genomic Insights of First ermB-Positive ST338-SCCmecVT/CC59 Taiwan Clone of Community-Associated Methicillin-Resistant Staphylococcus aureus in Poland.

We report the first Polish representative of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA), lukS/F-PV-positive, encoding the ermB gene, as a genetic determinant of constitutive resistance to macrolides, lincosamides, and streptogramin B antibiotics, cMLS-B. This is the first detection of the CA-MRSA strain responsible for nosocomial infection in the Warsaw Clinical Hospital. Resistance to β-lactams associates with a composite genetic element, SCCmec cassette type VT (5C2&5). We assigned the strain to sequence type ST338 (single-locus variant of ST59), clonal complex CC59, spa-type t437, and agr-type I. Genomic-based comparison was designated SO574/12 as an international Taiwan clone, which has been so far described mainly in the Asia-Pacific region. The ermB gene locates on the chromosome within the 14,690 bp mobile element structure, i.e., the MESPM1-like structure, which also encodes aminoglycoside- and streptothricin-resistance genes. The MESPM1-like structure is a composite transposon containing Tn551, flanked by direct repeats of IS1216V insertion sequences, which probably originates from Enterococcus. The ermB is preceded by the 273 bp regulatory region that contains the regulatory 84 bp ermBL ORF, encoding the 27 amino acid leader peptides. The latest research suggests that a new leader peptide, ermBL2, also exists in the ermB regulatory region. Therefore, the detailed function of ermBL2 requires further investigations.

Safety evaluation of enterococci isolated from raw milk and artisanal cheeses made in Slovenia and Serbia.

Enterococci represent a significant part of the non-starter LAB microbiota of artisanal cheeses produced mainly from raw milk. Common approaches to safety evaluation of enterococci isolates include assessment of antimicrobial resistance and virulence potential. Hence, a collection of 47 (n = 22, Serbia; n = 25, Slovenia) dairy enterococcal isolates, of which E. faecalis (n = 28), E. faecium (n = 11), E. durans (n = 5), E. casseliflavus (n = 2), and E. gallinarum (n = 1), was analyzed. The susceptibility to 12 antimicrobials was tested using a broth microdilution method, and the presence of the selected antimicrobial resistance and virulence genes was investigated using PCR. Isolates were resistant to tetracycline (TET) (25.5%), erythromycin (ERY) (17.0%), gentamycin and chloramphenicol (CHL) (∼6%). No resistance to ampicillin (AMP), ciprofloxacin (CIP), daptomycin (DAP), linezolid (LZD), teicoplanin (TEI), tigecycline (TGC) and vancomycin (VAN) was detected. Among all the resistance determinants analyzed, ermB gene was detected most frequently. All 10 virulence genes analyzed were detected with a distribution of cpd (72.3%), cob and ccf (70.2%), gelE (68.1%), hyl (59.6%), agg (53.2%) and esp (46.8%). The genes encoding cytolysin (cylA, cylM and cylB) were amplified to a lesser extent (21.3%, 21.3% and 12.8%, respectively). However, due to the limited number of enterococci isolates analyzed in the present study, further studies are still required in order to better document the safety status of dairy enterococci.

Phenotypic and Genotypic Characterization of Macrolide, Lincosamide and Streptogramin B Resistance among Clinical Methicillin-Resistant Staphylococcus aureus Isolates in Chile.

Macrolides, lincosamides, and type B streptogramins (MLSB) are important therapeutic options to treat methicillin-resistant Staphylococcus aureus (MRSA) infections; however, resistance to these antibiotics has been emerging. In Chile, data on the MLSB resistance phenotypes are scarce in both community-(CA) and hospital-acquired (HA) MRSA isolates. Antimicrobial susceptibility to MLSB was determined for sixty-eight non-repetitive isolates of each HA-(32) and CA-MRSA (36). Detection of SCCmec elements, ermA, ermB, ermC, and msrA genes was performed by PCR. The predominant clones were SCCmec I-ST5 (HA-MRSA) and type IVc-ST8 (CA-MRSA). Most of the HA-MRSA isolates (97%) showed resistance to clindamycin, erythromycin, azithromycin, and clarithromycin. Among CA-MRSA isolates, 28% were resistant to erythromycin, azithromycin, and 25% to clarithromycin. All isolates were susceptible to linezolid, vancomycin, daptomycin and trimethoprim/sulfamethoxazole, and over 97% to rifampicin. The ermA gene was amplified in 88% of HA-MRSA and 17% of CA-MRSA isolates (p < 0.001). The ermC gene was detected in 6% of HA-SARM and none of CA-SARM isolates, whereas the msrA gene was only amplified in 22% of CA-MRSA (p < 0.005). Our results demonstrate the prevalence of the cMLSB resistance phenotype in all HA-MRSA isolates in Chile, with the ermA being the predominant gene identified among these isolates.

Vertical distribution of antibiotics and antibiotic resistance genes in a representative municipal solid waste landfill, China

The vertical distribution of sulfonamides (SAs), tetracyclines (TCs), macrolides (MLs), and their related antibiotic resistance genes (ARGs) were comprehensively investigated and characterized in a representative municipal solid waste (MSW) landfill in China. The total concentrations of target antibiotics in the MSW landfill were SAs > TCs > MLs. The abundances of mexF (10.78 ± 0.65 log10copies/g) and sul genes (9.15 ± 0.54 log10copies/g) were relatively high, while the tet genes (7.19 ± 0.77 log10copies/g) were the lowest. Both the abundance of antibiotics and genes fluctuated with landfill depth, and the ARGs of the same antibiotics were consistent with depth change. Intl1 and sul genes (sul1, sul2) were tightly connected, and a close relationship also existed between tet genes (tetM, tetQ) and MLs resistance genes (ermB, mefA). High-throughput sequencing showed the dominant genera were Sporosarcina (38%) and Thiobacillus (17%) at sampling points A and C, while the microbial community varied with depth increase at point B were Brevundimonas (20%), Sporosarcina (20%), Pseudomonas (24%), Lysobacter (28%), and Thioalkalimicrobium (14%), respectively. Network analysis further visualized the relationship among antibiotics, genes, and microbial communities and the results indicated the non-random connection among them and the possible host of the target gene. Even at 12.0 m below the landfill surface, the pollution of antibiotics resistance was still serious, which posed difficulties for subsequent landfill remediation and pollution control.

Development and clinical application of a rapid and visual loop-mediated isothermal amplification test for tetM gene in Clostridioides difficile strains cultured from feces

ObjectivesIn this study, we aimed to develop a rapid and visual loop-mediated isothermal amplification (LAMP) assay targeting the tetM gene in Clostridioides difficile strains cultured from feces. MethodsPrimers were designed to recognize the tetM gene in C. difficile by LAMP, using turbidity and visual detection. The sensitivity and specificity of LAMP primers were determined. In addition, we conducted both LAMP and polymerase chain reaction (PCR) for the tcdA, tcdB, cdtA, cdtB, ermB, and tetM genes in 300 toxigenic C. difficile strains cultured from feces. ResultsThe target DNA was amplified and visualized within 60 minutes at a temperature of 62°C. A total of 26 bacterial strains were found negative for tetM, which manifested high specificity of the primers. The detection limit of LAMP was 36.1 pg/µl, which was 100-fold more sensitive than PCR. The positive rate of tetM in toxigenic C. difficile strains cultured from feces was 93.3% by both LAMP and PCR. The proportion of toxin types in those C. difficile strains was 95.7% for A+B+CDT−, 4% for A−B+CDT−, and 0.3% for A+B+CDT+, respectively. ConclusionThis is the first study examining the tetM gene by LAMP in C. difficile strains cultured from feces. Its high specificity, sensitivity, and visual detection make the new assay a powerful diagnostic tool for rapid testing.

Discovery of Novel Lincomycin Derivatives Effective against Resistant Streptococcus pneumoniae and Streptococcus pyogenes Possessing ErmB Gene

Discovery of Novel Lincomycin Derivatives Effective against Resistant Streptococcus pneumoniae and Streptococcus pyogenes Possessing ErmB Gene

A solution to combat Erythromycin-resistant bacteria isolated from returned activated sludge in Salt Lake City, Utah

Treatments offered in clinics are now ineffective due to antibiotic resistance in particular bacterial strains. Plant-based antibiotics are in high demand in developing and developed countries; they are common medications that are simple to use, pose no environmental risks, have no adverse side effects, and are competitively priced. This study aimed to screen plant-based medicine against Erythromycin-resistant bacteria such as E. coli, Enterobacter, Enterococcus, and E. Faecalis. The disk diffusion method and the agar well diffusion method were used to determine the zone of inhibition of Coriandrum Sativum L. (coriander), Zingiber officinale (ginger), and Spinacia oleracea (spinach). Minimum inhibitory concentration was evaluated via UV-visible Spectrophotometry at 600nm, while Polymerase Chain Reaction and Electrophoresis identified genomic activity for ErmB. Coriander was found to be the most effective against Erythromycin-resistant bacteria, and ErmB genes were found in almost all of the isolates.&#x0D; Keywords: Coriandrum Sativum, Zingiber officinale, Spinacia oleracea, Antimicrobial activity, and Erythromycin

Multidrug-resistant Bacterial Isolates in Bovine Subclinical Mastitis from Southern Rajasthan

Background: In the present study two hundred milk samples were collected from cows and buffaloes with no history of clinical mastitis in the ongoing lactation, from the Sirohi district of Southern Rajasthan. Methods: The pooled sample (collected from each quarter) and examined for the status of subclinical mastitis by Modified California mastitis test and Somatic cell count respectively. Positive samples were further investigated for isolation and identification of the major mastitis-causing pathogens: S. aureus, predominant Streptococcal species and E. coli for assessing antimicrobial resistance models in southern Rajasthan. Result: The results of the current study indicate high levels of multi-drug antibiotic resistance among bacteria that commonly cause mastitis, particularly ampicillin, penicillin, tetracycline, erythromycin and methicillin. However, the highest sensitivity was conferred to ceftriaxone, gentamicin, and co-trimoxazole, suggestive of judicious use of these antibiotics in the treatment of bovine mastitis. Concurrent implementation of gradient PCR indicated the presence of mecA and blaZ genes in 51.9% and 81.4% of S. aureus isolates, respectively. Meanwhile, 56.6% of the streptococcal isolate contained the tetracycline-conferring tetM gene and none of the streptococci contained the ermB gene. The 92.3% E. coli isolates contained the tetA gene and the tetB gene for tetracycline resistance.

Multiplex real-time PCR using SYBR Green: Unspecific intercalating dye to detect antimicrobial resistance genes of Streptococcus pneumoniae in cerebrospinal fluid.

Meningitis caused by Streptococcus pneumoniae is still a disease of great impact on Public health, which requires immediate diagnosis and treatment. However, the culture of clinical specimens is often negative and antibiotic susceptibility testing (AST) must be performed with isolated strains. Multiplex real-time polymerase chain reaction (qPCR) has high sensitivity and specificity, produces faster results to identify the pathogen, and it can also be an important tool to identify resistance antibiotic genes earlier than AST, especially in the absence of an isolated strain. This study developed a multiplex qPCR assay, using SYBR Green as a nonspecific dye, to detect antibiotic resistance genes to predict pneumococcal susceptibility/resistance in cerebrospinal fluid (CSF) samples from meningitis patients. From 2017 to 2020, CSF samples were cultured and analyzed by qPCR to detect the main three bacteria causing meningitis. Isolated and reference strains were applied in SYBR Green qPCR multiplex to detect pbp2b, ermB, and mef genes, and the results were compared with the AST. Pneumococcal-positive CSF samples (lytA-positive gene) without isolated strains were also tested to evaluate the antimicrobial susceptibility profile in the region from 2014 to 2020. From the received 873 CSF samples; 263 were cultivated, 149 were lytA-positive in the qPCR, and 25 produced viable isolated pneumococci strains, which were evaluated by AST. Melting temperature for each gene and the acceptance criteria were determined (pbp2b: 78.24-79.86; ermB: 80.88-82.56; mef: 74.85-76.34 ºC). A total of 48/51 strains presented a genetic profile in agreement with the AST results. Resistant strains to erythromycin and clindamycin were ermB-positive, and two were also mef-positive, indicating both resistance mechanisms were present. In the retrospective study of the genetic profile of resistance, 82 lytA-positive CSF samples plus 4 strains were applied in the SYBR Green qPCR multiplex: 51% of samples presented the wild genotype (pbp2b positive and ermB/mef negative); 15% were negative for all the three evaluated, indicating pneumococci resistant to penicillin; and 17% represented the multidrug-resistant pneumococci (pbp2b negative and ermB positive or pbp2b negative and ermB and mef positive). Therefore, SYBR Green qPCR multiplex proved to be a reliable tool to identify resistance genes in S. pneumoniae and would be less expensive than multiplex qPCR using specific probes. This could be easily introduced into the routine of diagnostic laboratories and provide a strong presumption of pneumococcal resistance, especially in the absence of isolated strains.

Antimicrobial Activity of the Iron-Chelator, DIBI, against Multidrug-Resistant Canine Methicillin-Susceptible Staphylococcus pseudintermedius: A Preliminary Study of Four Clinical Strains.

Staphylococcus pseudintermedius is an important opportunistic pathogen causing various infections in dogs. Furthermore, it is an emerging zoonotic agent and both multidrug-resistant methicillin-resistant S. pseudintermedius (MRSP) as well as methicillin-susceptible (MSSP) strains represent an important therapeutic challenge to veterinary medicine and pose a potential threat to human health. We tested representative S. pseudintermedius clinical strains from dogs suffering from otitis externa for their susceptibilities to a panel of 17 antimicrobials compared to DIBI. DIBI, unlike antibiotics, is a novel water-soluble hydroxypyridinone-containing iron-chelating agent that deprives microbes of growth-essential iron and has been previously shown to inhibit methicillin-resistant Staphylococcus aureus (MRSA). We also characterised the strains according to whether they harbour key antibiotic resistance genes. The strains each displayed multiple antimicrobial resistance patterns; all were negative for the mecA gene and possessed the tetK and tetM genes, but they varied as to their possession of the ermB gene. However, all the isolates had similar susceptibility to DIBI with low MICs (2 µg/mL or 0.2 µM). Because the four MSSPs were equally susceptible to DIBI, subject to confirmation with additional strains, this could provide a potential non-antibiotic, anti-infective alternative approach for the treatment of antimicrobial-resistant canine S. pseudintermedius otitis.

Clinical and Genomic Investigation of an International Ceftriaxone- and Azithromycin-Resistant Shigella sonnei Cluster among Men Who Have Sex with Men, Montréal, Canada 2017-2019.

ABSTRACTMultidrug-resistant (MDR) Shigella sonnei have become prevalent among men who have sex with men and have become a global public health concern. From June 2017 to April 2019, 32 men were infected with MDR S. sonnei acquired locally, in Montréal, which was suggestive of an outbreak. Antimicrobial susceptibility testing, whole-genome sequencing (WGS), phylogenetic analysis, antimicrobial resistance and virulence characterization, and association to international clusters were performed. The outbreak strain was ceftriaxone- and azithromycin-resistant due to the acquisition of blaCTX-M-27, and mphA and ermB genes, respectively, with reduced susceptibility to ciprofloxacin due to a single point mutation (gyrA S83L). One out of 27 patients treated with a fluoroquinolone experienced microbiological failure. Epidemiological evidence first supported by a rare unique MDR Shigella sonnei documented only in men in 2017 followed by similar pulsed-field gel electrophoresis profiles was confirmed by WGS. A core genome high-quality single-nucleotide variant (hqSNV)-based phylogeny found a median of 6 hqSNV differences among isolates. Virulence gene content was investigated, but no Shiga toxins were detected. An international cluster of highly related isolates was identified (PDS000019750.208) and belonged to the 3.7.29.1.4.1 S. sonnei genotype (Global III VN2.KH1.Aus). Genomic analysis revealed that this Montréal cluster was connected to other documented outbreaks in Australia, the United States, and the United Kingdom. This study highlights the urgent need for public health measures to focus on the prevention and the early detection of S. sonnei, since global transmission patterns of MDR strains is concerning and few antimicrobial treatment options are available.IMPORTANCE Shigella sonnei, an important foodborne pathogen, recently became a frequent sexually transmitted agent involved in large and persistent outbreaks globally among men who have sex with men. Most strains also harbor several multidrug-resistant (MDR) determinants of particular concern. This study characterizes an outbreak strain at the source of an important MDR cluster identified in Montréal in 2017. Associations were made to many high-profile international outbreaks, and the causative S. sonnei lineage of these clusters was identified, which was not evident in past reports. The worldwide occurrence of this strain is of concern since treatment with antimicrobials like ceftriaxone and azithromycin may not be effective, and rare microbiological failures have been documented in patients treated with ciprofloxacin. Our investigation highlights the threats of Shigella spp. infection and the necessity for antimicrobial susceptibility monitoring in order to mitigate S. sonnei’s impact on public health and to avoid transmission to other at-risk communities.

Human enterococcal isolates as reservoirs for macrolide-lincosamide-streptogramin and other resistance genes.

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

Antimicrobial Resistance of Secondary Bacterial Infections in Patients Hospitalized with Covid-19

Coronavirus disease 2019 (COVID-19) is an emerging serious global health problem. It has been recognised for a considerable time-period, that viral respiratory infections predispose patients to bacterial infections, and that these co-infections have a worse outcome than either infection on its own. This study was carried out on 100 samples of sputum from COVID-19 patients. During the laboratory diagnosis, 156 bacterial isolates were obtained from the positive samples . The Gram-positive bacteria isolates included Strptococcus pneumonia 64(40%) Streptococcus pyogenes, 7 (4%), Streptococcus mitis 1 (1%) Streptococcus mutus 1(1%) , Streptococcus parasanguinis 1(1%) Staphylococcus eidermidis 10(6%), staphylococcus aureus 4 (2%), Micrococcus lutus,1 (1%). Whereas, Gram-negtive bacteria included Pseudomonas aeruginosa 9 (6%), E coli 10 (6%), Serratia marcescens, 3 (2%), Klebsiella pneumonia 31 (19%), H.influenzae 10 (6%) Acinetobacter baumannii , 4 (2%) .The isolates varied in their response against the antibiotics ; and Gram positive bacteria were significantly (p&lt;0.05) more sensitive to the antibiotic then Gram negative. To detect the Streptococcus pneumonia isolates , the house keeping Eno genes was screened. Results showed that all the isolates, had Eno gene (100%). Furthermore, This study was carried out in order to detect tet-L and ermB gene in 10 S.pneumonia isolates . genes were Results showed that all the isolates, numbering 10, had both tetL and ermB genes(100%). Keywords : Bacteria, Co-infections, COVID-19, Eno gene, tet-L gene , ermB gene

Antimicrobial resistance and virulence traits of Enterococcus faecium isolated from clinical bovine mastitis in Ningxia, China

This study was conducted to determine the antimicrobial resistance and virulence traits of 32 Enterococcus faecium isolates from clinical bovine mastitis cases in Ningxia Province, China. In total, 32 E. faecium isolates were taken from subclinical bovine mastitis on the basis of morphological characterization and biochemical testing, and screened for antimicrobial susceptibility. The virulence genes of the isolates were studied using polymerase chain reaction (PCR). The disc diffusion assay revealed a high occurrence of resistance against tetracycline (78.1%) and erythromycin (68.8%) in the E. faecium isolates. However, all tested E. faecium were susceptible to linezolid and vancomycin. Moreover, all E. faecium isolates harbored the erythromycin-resistant genes ermA, ermB and ermC, as well as the tetracycline-resistant genes tetK, tetL and tetM. Furthermore, all E. faecium isolates carried more than 3 of the tested virulence genes. The presence of agg (100%), cpd (100%), efaA (100%), gelE (93.4%), and esp (75.0%) was found most frequently in all the tested isolates. These findings are useful for making appropriate antimicrobial choices and developing antivirulence therapies for subclinical bovine mastitis caused by E. faecium in Ningxia Province, China.

MULTIDRUG-RESISTANT Salmonella sp. ISOLATED FROM SEVERAL CHICKEN FARMS IN WEST JAVA, INDONESIA

This study was aimed at isolating and identifying Salmonella sp. and then conducting an antibiotics susceptibility test in order to detect resistant genes. One hundred and five chicken cloaca swab samples were used in this study. 30 samples were taken from a layer farm in Bogor, 45 from a broiler farm in Sukabumi and 30 from a broiler farm in Cianjur. In order to isolate and identify the bacteria, a tetrathionate broth was used, which was then cultured in a Salmonella-Shigella agar, and finally a Gram stain and biochemical test was conducted. To confirm the presence of Salmonella sp., a pair of primers were used for the polymerase chain reaction (PCR) method to determine the presence of the invA gene.. An antibiotics susceptibility test was used with the Kirby-Bauer disk diffusion method. Nine antibiotics were used in this study. Each primer pair was used for the detection of tetA, blaTEM, aac(3)-IV, gyrA and ermB genes, and for genes encoding antibiotic resistance a PCR test was used. Eight (7.6%) Salmonella sp. were isolated in this study. All isolates showed positive results with PCR confirmation. The results of the antibiotics susceptibility test showed that Salmonella sp. isolates were resistant to tetracycline (75%), oxytetracycline (75%), amphicillin (75%), gentamycin (12.5%), nalidixic acid (100%), ciprofloxacin (12.5%), enrofloxacin (0%), erythromycin (100%), and chloramphenicol (0%). The distribution of antibiotic resistance genes in Salmonella sp. were tetA (33.3%), blaTEM (100%), aac(3)-IV (0%), gyrA (100%) and ermB (0%) positive. In conclusion, Salmonella sp. was isolated. All isolates showed positive results in the PCR confirmation. Salmonella sp. isolates were resistant to tetracycline, oxytetracycline, amphicillin, gentamycin, nalidixic acid, ciprofloxacin, and erythromycin. Only the tetA, blaTEM, and gyrA genes were detected in Salmonella sp. isolates.

Species distribution and genes encoding antimicrobial resistance in enterococcus spp. isolates from rabbits residing in diverse ecosystems: A new reservoir of linezolid and vancomycin resistance.

Worldwide, studies regarding antimicrobial resistance in rabbits are scarce. In addition, it seems that rearing conditions have important impact on emergence and spread of antimicrobial-resistant bacteria. Thus, the authors sought to (1) assess the role of rabbits residing across diverse ecosystems as potential reservoirs of antimicrobial-resistant enterococci and (2) investigate the genetic background of detected resistances. Faecal samples from 60 healthy farmed rabbits (one farm), 35 laboratory rabbits and 31 wild rabbits were analysed. Overall, 97 enterococci isolates were accumulated, as follows: 44 E. faecium, 37 E. faecalis, 7 E. gallinarum, 5 E. durans and 4 E. avium. E. faecalis isolates were statistically associated with farm rabbits and wild rabbits (p<0.05). High rates of resistance were observed for tetracycline (60.8%; tetM [n=48; 81.3%], tetO [n=7; 11.8%] and tetL [n=1; 1.7%]), erythromycin (43.3%; msr(A) [n=14; 33.3%] and ermB [n=13; 31%]), ampicillin (29.9%), streptomycin (26.8%; ant(6)-Ia [n=3, 11.5%]) and vancomycin (21.6%; vanA [one E. faecium+one E. faecalis; 9.5%]). Low frequencies of resistance were observed for teicoplanin (9.2%), linezolid (8.2%), ciprofloxacin (7.2%) and gentamicin (1%; aac(6')-Ie-aph(2″)-Ia). Resistance to ampicillin and vancomycin was associated with laboratory rabbits (p<0.05). Int-Tn (Tn916/1545) was detected in 27 (27.8%) isolates, of which 10 isolates co-harboured tetM and ermB genes, while 16 comprised tetM. Findings indicate that clinically relevant enterococci species isolated from rabbits are frequently resistant to antimicrobials and harbour a range of genes associated with the Tn916/1545 family. This study highlights the high rates of antimicrobial-resistant enterococci from rabbits and the occurrence of both vancomycin- and linezolid-resistant isolates, potentially representing a very serious threat to human and animal health.

Serotype distribution and antimicrobial resistance of invasive Streptococcus pneumoniae isolates among Croatian adults during a fifteen-year period (2005-2019)

AimTo assess serotype distribution, antibiotic resistance, and vaccine coverage against Streptococcus pneumoniae causing invasive infections in Croatian adults from 2005 to 2019.MethodsIn this retrospective study, invasive pneumococcal strains were collected through a microbiological laboratory network with country coverage >95%. Capsular typing was performed with the Quellung reaction. In vitro susceptibility testing was carried out according to the European Committee on Antimicrobial Susceptibility Testing guidelines. In macrolide-resistant isolates, the presence of ermB and mefA genes was evaluated.ResultsDuring the fifteen-year study period, 1123 invasive pneumococcal isolates were obtained. The most prevalent serotypes were 3, 14, 19A, 9V, 7F, and 23F, comprising 60% of all invasive pneumococcal isolates. Serotype 3 was the dominant serotype, with the highest prevalence in patients ≥65 years of age. Penicillin susceptibility, increased exposure was 18.6%, mostly associated with serotypes 14 and 19A. Resistance to penicillin was low (<1%). Macrolide resistance was 23%, mostly associated with serotypes 14, 19A, and 19F. The coverage with 13-valent conjugate vaccine (PCV13) and 23-valent polysaccharide vaccine (PPV23) was 80.2% and 93.6%, respectively.ConclusionsThe incidence of invasive pneumococcal disease in adults is highest in patients ≥65 years of age. Penicillin susceptibility, increased exposure and macrolide resistance were mostly associated with serotypes 14 and 19A. PCV13 and PPV23 provide very high serotype coverage. Future studies should evaluate the effects of the 10-valent vaccine, introduced in the Croatian National Immunization Program in June 2019, on serotype distribution and antibiotic resistance rates.