Bacitracin Susceptibility Research Articles

Skin infections caused by Streptococcus pyogenes after the Eid Al-Adha

Introduction: Streptococcus pyogenes is Gram-positive bacteria in chain form. Infections typically begin in the throat and skin, although they present with a many different clinical cases. This study aimed to present a different clinical manifestation related to S. pyogenes. Methods: In this study, swab samples were taken from patients who came to our center with skin lesions after Eid al-Adha. Identification was performed using conventional methods (bacitracin susceptibility test and PYR test) and VITEK 2 (bioMerieux, Marcy l’Etoile, France). Phylogenetic tree was generated using the Tamura Nei model of Neighbor-joining method in MEGA 6.0 program. Results: All of the isolated microorganisms were defined as S. pyogenes. Phylogenetic tree revealed that strains were highly related, but there were genetic differences between the strains. Conclusion: While Orf virus (Parapoxvirus) is the most common cause of skin lesions after Eid al-Adha, S. pyogenes was found to be the causative agent in our study. To our knowledge, cases of S. pyogenes skin infection due to religious practices have been reported for the first time.

Vertical Transmission, Risk Factors, and Antimicrobial Resistance Patterns of Group B Streptococcus among Mothers and Their Neonates in Southern Ethiopia.

Background Group B Streptococcus (GBS) contributes to maternal and neonatal morbidity and mortality by increasing intrauterine infection or vertical transmission at the time of birth. Despite many efforts to reduce the potential risk of vertical transmission, GBS remains the main cause of serious disease (neonatal sepsis, meningitis, and/or pneumonia) in vulnerable newborns during the first week of life. This study aimed to assess vertical transmission, risk factors, and antimicrobial resistance patterns of GBS among pregnant women and their neonates. Methods A facility-based cross-sectional study was conducted among mothers and their neonates from February to May 2021. A total of 201 pregnant women with their neonates participated in this study. A well-designed questionnaire was used to collect sociodemographic and clinical data. A vaginal swab from mother before delivery and neonatal nasal and ear canal swab samples were taken as soon as after delivery within 30 minutes. Vaginal swabs, neonatal ear canal, and nasal swabs were placed into Todd–Hewitt broth and incubated at 37°C for 18–24 hours at 35–37°C in 5% CO2 conditions and then subcultured on 5% sheep blood agar for 18–48 hours. Presumptive identification of GBS was made by morphology, Gram stain, catalase, and hemolytic activity on sheep blood agar plates. CAMP and bacitracin susceptibility tests were used as confirmatory tests for GBS. Data were analyzed using SPSS version 21. P value ≤0.05 was considered statistically significant. Results Vertical transmission rates of GBS (mother to neonates) were 11.9%. The prevalence of GBS among pregnant women and newborns was 24/201 (11.9%) (95% CI = 7.5–16.9) and 11/201 (5.5%) (95% CI = 2.5–9.0), respectively. The history of prolonged rupture of membranes (AOR = 3.5, CI = 2.2–18.8) and urinary tract infection (AOR = 2.9, CI = 1.7–16.3) were associated factors for maternal GBS colonization. Gestational age of <37 weeks (p=0.008), low birth weight of <2.5 kg (p=0.001), and maternal history of vaginal discharge (p=0.048) were associated factors for neonatal GBS colonization. Low antibiotic resistance was observed for erythromycin 8.6%, clindamycin 5.7%, and chloramphenicol 2.9%. Conclusion In this study, high vertical transmission (mother to neonates) rate was observed. The prevalence of vaginal GBS colonization of women at delivery was 11.9% and significantly associated with the history of prolonged rupture of membranes and urinary tract infections. Gestational age of <37 weeks, low birth weight of <2.5 kg, and maternal history of vaginal discharge were associated with neonatal GBS colonization. Hence, there is a need for antenatal culture-based GBS screening, risk factor-based interventions, and regular follow-up of drug resistance patterns for proper treatment and management of GBS.

Streptococcal Pharyngitis and Rheumatic Fever

Streptococcus pyogenes (Group A Streptococcus) causes a variety of diseases, from benign self-limiting infections of the skin or throat to lethal infections of soft tissue accompanied by multi-organ failure. GAS is one of significant species among Gram-positive pathogens which is responsible for several suppurative infections and non-suppurative sequelae. They also cause pharyngitis, streptococcal toxic shock syndrome (STSS), necrotizing fasciitis and other diseases. Currently, global burden of RF / RHD is undervalued. In 2010, RF and RHD were estimated as 15.6 million cases and deaths around 200,000 annually. Laboratory diagnosis includes cultural techniques, serology, PYR test, Bacitracin susceptibility test and antibiotic resistance testing helps in differentiating the Streptococcus pyogenes from other groups of Streptococci. Most of the Acute Rheumatic Fever cases gets missed or does not present in the initial stage rather it has been developed into advanced Rheumatic Heart Disease condition. Modified Jones criteria in 2015 will be helpful especially to the low risk population as it is challenging because of limited access to primary health care, diagnosis of streptococcal disease. In addition to this revised criteria, diagnosis still relies on clinical diagnostic algorithm. Vaccines based on M protein and T antigens are continuing to evolve with different results. Ongoing vaccine development is still challenging for the GAS research community, it will make a positive and lasting impact on the peoples globally.

Prevalence and the risk factors for the carriage of beta-haemolytic streptococci among women visiting a tertiary care hospital in South India

Streptococci are gram positive cocci arranged in chains and are part of normal flora of humans and animals. The present study is carried out to determine the prevalence and risk factors for the carriage of beta-haemolytic streptococci (BHS) among women visiting Dr. VRK Women’s Teaching Hospital &amp; Research Centre, Hyderabad. Vaginal swabs were collected from 250 patients attending outpatient department (OPD) of Dr. VRK Women’s Teaching hospital. Swabs were inoculated onto 5% sheep blood agar plates and incubated for 24 h at 37°C in a candle jar. BHS isolates were phenotypically identified by standard microbiological techniques, all the isolates presumptively identified as BHS were tested for Bacitracin susceptibility. Sensitive isolates were presumptively identified as GAS and resistant isolates were identified as non-group A BHS (NGABHS). Presumptively identified GAS &amp; NGABHS isolates were serogrouped by Lancefield grouping using a commercially available latex agglutination test. BHS were isolated from 12.4% of samples. As many as 12 BHS isolates were identified as GAS and 19 were identified as NGABHS. Ten of nineteen were identified as group B (GBS), 4 (12.9%) were identified as group C (GCS) and 5 (16.12%) were identified as group G (GGS). Among six clinical groups, the prevalence of GAS is highest i.e. 7.5% in female patients visiting Gynaecology OPD with history of white discharge. Prevalence of NGABHS was more among post insertion (18%) IUCD group compared to pre insertion (8%) IUCD group. GBS were isolated from 7% of samples from IUCD group and 4% of samples from prostitutes.This study reports the prevalence of BHS among women visiting a tertiary care hospital in Hyderabad. This study also identified certain risk factors such as IUCD usage and working as a FSW are associated with the increased prevalence of NGABHS especially GBS.

Prevalence and antibiotic susceptibility of Streptococcus pyogenes isolated from pyoderma in a tertiary care hospital, Hyderabad, South India

Pyoderma is a common acute superficial bacterial skin infection which is highly contagious. In the great majority of cases, pyoderma is caused by , , or both. The present study was carried out to determine the prevalence and antibiotic susceptibility of isolated from pyoderma in Dr. VRK Women’s Teaching hospital.Swabs or pus samples were collected from 250 patients attending Dermatology, outpatient department (OPD) of Dr. VRK Women’s Teaching hospital. Samples were inoculated onto 5% sheep blood agar plates and incubated for 24 h at 37°C in a candle jar. BHS isolates were phenotypically identified by standard microbiological techniques, all the isolates presumptively identified as BHS were tested for Bacitracin susceptibility. Presumptive identification of a strain as a Group A Streptococcus (GAS) was also made by PYRase test. Presumptively identified GAS isolates were serogrouped by Lancefield grouping using a commercially available latex agglutination test. isolates were subjected to antimicrobial susceptibility testing by Kirby-Bauer disc diffusion method.BHS were isolated from 30% of samples. Prevalence of BHS was more among 0-10 years age group (38%). BHS were isolated more frequently from males (38.8%). were isolated from 52 (20.8%) samples. All 52 isolates were found to be susceptible to Penicillin G, amoxicillin, ceftriaxone, azithromycin and vancomycin. Erythromycin and clindamycin showed good activity with sensitivity rates of 92.3% &amp; 96.1%, respectively. Resistance to tetracycline (59.6%) and chloramphenicol (23.1%) was commonly seen in .This study reports the prevalence and antibiotic susceptibility of isolated from pyoderma in Dr. VRK Women’s Teaching hospital. Results of this study suggests the peak incidence of pyoderma in children aged 0 to 10 years and male preponderance. Our study also reports high prevalence of tetracycline and chloramphenicol resistance in .

Molecular epidemiological characteristics of Streptococcus pyogenes causing scarlet fever and angina in children

Objective To investigate the antimicrobial resistance, macrolide-resistance genes, virulence genes and emm types in Streptococcus pyogenes isolates. Methods A total of 247 oropharyngeal swab specimens were collected from pediatric outpatients (aged 2-11 years) who were clinically diagnosed as scarlet fever in Children′s Hospital of Fudan University from January to December, 2018. These specimens were timely sent to the Microbiology Laboratory for isolation and identification of Streptococcus pyogenes strains were isolate after culturing and identified with bacitracin susceptibility test. Moreover, the diameter of bacitracin inhibition zone was measured by vernier caliper. Their susceptibility to seven antibiotics, including erythromycin, clarithromycin, clindamycin, ampicillin, ceftriaxone, norfloxacin and chloramphenicol, were measured using KB method. Macrolide-resistance genes (mefA, ermA, ermB and Tn916 transposon) and virulence genes (speA, speB, speC, speG, speH, speI, speJ and speK) were detected by PCR. Amplification and sequencing of emm gene were conducted according to the protocol in the website of Centre for Disease Control and Prevention (CDC). Results A total of 86 strains of Streptococcus pyogenes were isolated from the 247 specimens. Their resistance rates to erythromycin, clarithromycin and clindamycin were 89.5%, 95.3% and 96.5%, respectively. However, these isolates showed high susceptibility to ampicillin (100.0%), ceftriaxone (100.0%), norfloxacin (90.7%) and chloramphenicol (95.3%). The positive rates of mefA, ermA, ermB and Tn916 genes were 20.9%, 24.4%, 98.8% and 97.7%. There was significant difference in the mefA-carrying rates between patients with scarlet fever and angina. The positive rates of virulence genes of speA, speB, speC, speG, speH, speI, speJ, speK, speL and speM were 8.1%, 100.0%, 95.3%, 100.0%, 80.2%, 90.7%, 10.5%, 100.0%, 5.8% and 5.8%. Seven emm types were identified and the predominant types were emm12.0 (75.6%), emm12.19 (9.3%) and emm1.0 (8.1%). The diameter of bacitracin inhibition zone was smaller in isolates of emm1.0 type than in emm12.0 type strains. The profile of virulence genes varied in the strains of different emm types. All types of strains carried speB, speG and speK genes. The isolates of emm1.0 type carried speA virulence gene, while speB, speC, speG, speH, speI and speK genes were more often identified in emm12.0 type isolates. Conclusions This study showed that emm types were associated with the profile of virulence genes and the diameter of bacitracin inhibition zone. It was recommended that the diameter of bacitracin inhibition zone should be measured in bacitracin inhibition susceptibility test apart from only observing the formation of inhibition zone. Key words: Streptococcus pyogenes; Antimicrobial resistance; Virulence gene; emm type; Bacitracin susceptibility test

Bacitracin resistance and enhanced virulence of Streptococcus suis via a novel efflux pump

BackgroundStreptococcus suis is a prominent pathogen causing septicemia and meningitis in swine and humans. Bacitracin is used widely as a growth promoter in animal feed and to control the spread of necrotic enteritis in most developing countries. This study aimed to characterize a novel membrane transporter module Sst comprising SstE, SstF, and SstG for bacitracin resistance.ResultsComparative genomics and protein homology analysis found a potential efflux pump SstFEG encoded upstream of well-known bacitracin-resistance genes bceAB and bceRS. A four-fold decrease in bacitracin susceptibility was observed in sstFEG deletion mutant comparing with S. suis wildtype strain CZ130302. Further studies indicated that the bacitracin tolerance mediated by SstFEG is not only independent of the BceAB transporter, but also regulated by the two-component system BceSR. Given that SstFEG are harbored by almost all virulent strains, but not in the avirulent strains, we managed to explore its potential role in bacterial pathogencity. Indeed, our results showed that SstFEG is involved in S. suis colonization and virulence in animal infection model by its potential competitive survival advantage against host bactericidal effect.ConclusionTo our knowledge, this is the first study to functionally characterize the bacitracin efflux pump in S. suis to provide evidence regarding the important roles of the novel ABC transporter system SstFEG with respect to drug resistance and virulence.

An ABC Transporter of Streptococcus pneumoniae Involved in Susceptibility to Vancoresmycin and Bacitracin

Vancoresmycin is a novel tetramic acid antibiotic, probably interfering with functions of the cytoplasmic membrane. To investigate its mode of action, mutants of Streptococcus pneumoniae exhibiting reduced susceptibility to vancoresmycin were isolated at a low frequency. Four of them were further examined and showed similar pleiotropic phenotypes, including reduced growth rate, early autolysis, and chain formation. In one mutant, the level of transcripts from a single locus encoding the potential ABC transporter Spr0812-Spr0813 was increased sixfold. The corresponding DNA sequence revealed a nonsense mutation (C1744T) in spr0813, leading to the formation of a truncated permease lacking 2 of the 10 predicted transmembrane helices. As demonstrated by deletion and transformation analysis and reconstructing the spr0813(C1744T) mutation in the wild-type background, this nucleotide exchange was sufficient to cause reduced susceptibility to vancoresmycin and higher amounts of spr0812-spr0813 mRNA. Mapping and reporter assays of the cognate promoter P(abc) showed that spr0812 and spr0813 are cotranscribed with a preceding small gene and that the spr0813(C1744T) mutation does not affect the activity of P(abc). Due to the similarity of Spr0812-Spr0813 to bacitracin transporters of Streptococcus mutans and Bacillus spp., the bacitracin susceptibility of spr0813 mutants was examined. Both the spr0813(C1744T) nonsense mutation and the deletion of the transporter genes led to a clearly increased sensitivity to bacitracin. Thus, the intact transporter is required for intrinsic resistance to bacitracin, whereas reduced vancoresmycin susceptibility is mediated by the truncated permease.

Bacitracin-resistant clone of Streptococcus pyogenes isolated from pharyngitis patients in Belgium.

We report 16 bacitracin-resistant Streptococcus pyogenes isolates recovered from pharyngitis patients in Belgium, 14 of which belonged to a particular emm type (emm28). All 16 isolates were constitutively resistant to macrolides and carried erm(B). The emergence of a bacitracin-resistant S. pyogenes clone raises questions about the continued reliability of bacitracin susceptibility testing for S. pyogenes identification.

Use of second-line biochemical and susceptibility tests for the differential identification of coryneform bacteria

Objective To investigate which ‘second-line' biochemical and susceptibility tests could be used for the differential identification of coryneform bacteria ( Actinomyces, Arcanobacterium, Brevibacterium, Corynebacterium, Dermabacter, Rothia, Turicella ). Methods Tests used for the diagnosis of certain Gram-negative rods and nocardiforms: growth in 10% NaCl; production of β-galactosidase, arginine dihydrolase, ornithine and lysine decarboxylase, γ-glutamylaminopeptidase, and methanethiol; hydrolysis of tributyrin, casein, tyrosine, and starch; assimilation of carbohydrates in the APl 20 NE system; gas-liquid chromatography for end products of glucose metabolism, and susceptibility to antimicrobials used for diagnostic purposes. Results Some tests proved useful for the differentiation within the genus Corynebacterium (e.g. growth in 10% NaCl, tyrosine hydrolysis, furazolidine and O/129 susceptibility), while others differentiated only between genera (e.g. methanethiol formation, bacitracin susceptibility). Conclusion Second-line tests can be of use for the differential identification of coryneform bacteria.

The family pet as an unlikely source of group A beta-hemolytic streptococcal infection in humans

This study examines the possibility of the family pet serving as a reservoir for group A beta-hemolytic streptococcal infections in humans. We obtained oropharyngeal cultures from children with acute pharyngitis and concurrent oropharyngeal cultures from their household pets. Children with culture-proved group A beta-hemolytic streptococcal pharyngitis were detected in 26 of 42 households surveyed. Oropharyngeal cultures were also collected from a group of children without pharyngitis and their pets. Additionally 149 dogs and cats from a local veterinary hospital were cultured from 371 body sites including the oropharynx, axilla and vagina. All beta-hemolytic bacterial isolates were identified by colonial and microscopic morphology, catalase and pyrrolidonylarylamidase production, bacitracin susceptibility and serogrouping. No group A beta-hemolytic streptococci were recovered from any of the body sites surveyed from a total of 230 animals. Based on these findings, the family pet seems to be an unlikely reservoir for group A beta-hemolytic streptococci.

Use of Lysostaphin and Bacitracin Susceptibility for Routine Presumptive Identification of Staphylococci of Bovine Origin

A simple agar plate system based on susceptibility to lysostaphin and bacitracin to differentiate between staphylococci and micrococci is described. The system also uses susceptibility to novobiocin to aid in identification to species level of staphylococci. Growth from the agar plate can be used to prepare inocula for inoculation of rapid identification systems.

Evaluation of the Strep-A-Chek technique for presumptive identification of group A β-hemolytic streptococci and group D enterococci

Strep-A-Check (E-Y Laboratories, San Mateo, CA) is a 15-min chromogenic test for a species-specific aminopeptidase that could replace testing bacitracin susceptibility for presumptive identification of group A streptococci as well as 6.5% NaCl agar tolerance for presumptive identification of enterococcal streptococci, with a time savings of 24 hr. Recent clinical streptococcal isolates (n = 341), identified by conventional biochemical and serologic techniques, were used to evaluate the 15-min Strep-A-Chek test. Among the beta-hemolytic streptococci (176 group A, 43 group B, 8 group C, 9 group F, and 9 group G), Strep-A-Chek was 100% accurate. Among the non-beta-hemolytic streptococci, 100% of 52 group D enterococci, 100% of 30 viridans streptococci, and 100% of 14 group D nonenterococci were correctly identified by Strep-A-Chek. Strep-A-Chek is an extremely rapid and reliable test for presumptive identification of group A and enterococcal streptococci.

Variations in bacitracin susceptibility observed in Staphylococcus and Micrococcus species.

Bacitracin susceptibility was evaluated as a laboratory method to differentiate staphylococci from micrococci. A total of 317 staphylococcal isolates and 108 micrococcal isolates were each tested for susceptibility to bacitracin by a disk-diffusion method using disks of three different potencies (0.04, 2.0, and 10.0 U) and a broth dilution method to obtain MICs. When a growth inhibition zone diameter breakpoint of greater than 10 mm was used to establish susceptibility with a 0.04-U disk, all micrococci were bacitracin susceptible and 94.6% of the staphylococci were resistant. Testing with disks of higher potency did not improve the specificity of the disk-diffusion method.

Incidence and characterization of beta-hemolytic Streptococcus milleri and differentiation from S. pyogenes (group A), S. equisimilis (group C), and large-colony group G streptococci.

The biochemical characteristics of 172 clinical isolates of group A, C, F, or G or "nongroupable" beta-hemolytic streptococci were examined. Among these isolates, 91 were identified as beta-hemolytic strains of Streptococcus milleri. The remaining isolates included 20 Streptococcus pyogenes, 21 Streptococcus equisimilis, 37 large-colony group G streptococci, and 3 unidentified nongroupable isolates. A majority (84%) of the S. milleri strains possessed Lancefield group antigen (3 A, 27 C, 41 F, and 5 G), whereas 15 S. milleri strains (16%) were nongroupable. Serological tests did not differentiate S. milleri isolates with group A, C, or G antigen from S. pyogenes (group A), S. equisimilis (group C), or large-colony group G streptococci. Biochemical tests which were found useful for differentiation included the Voges-Proskauer test, hydrolysis of pyroglutamic acid and beta-D-glucuronide, bacitracin susceptibility, and acid production from ribose. S. milleri represented 56% of the group C, 100% of the group F, and 83% of the nongroupable beta-hemolytic streptococci isolated in our clinical laboratory, whereas the incidence of S. milleri among group A and group G streptococci was estimated to be low. The role of beta-hemolytic S. milleri as a cause of human infection remains obscured by the failure to routinely differentiate S. milleri from other beta-hemolytic streptococci.

Comparison of fluorescent antibody, bacitracin susceptibility, latex agglutination, coagglutination, and API 20S for identifying group A streptococci

A total of 200 beta-hemolytic streptococci, isolated from clinical specimens submitted to our laboratory, were identified as group A versus non-A using the fluorescent antibody technique (FA), bacitracin susceptibility (BBL, Difco, and Raven disks), SeroSTAT, Streptex, Phadebact, and the API 20S system. Of the 122 group A isolates, all methods except SeroSTAT and Phadebact yielded 92-99% agreement when compared with the Lancefield precipitin test. Phadebact yielded an 84% agreement and SeroSTAT changed from 83 to 98% after trypsinization. Numerous false positives were obtained and only FA (91%) and API 20S (96%) yielded better than 90% agreement on non-A identification when compared with the Lancefield test. The most false positives were obtained (45%) using the SeroSTAT reagents. Considering accuracy, our data suggests the FA technique to be the method of choice for identifying group A streptococci.

Presumptive identification of streptococci with a new test system.

A test is described that could replace bacitracin susceptibility for presumptive identification of group A streptococci as well as 6.5% NaCl agar tolerance for presumptive identification of enterococcal streptococci. The L-pyrrolidonyl-beta-naphthylamide test, based on hydrolysis of pyrrolidonyl-beta-naphthylamide, was used in conjunction with the CAMP and bile-esculin tests to presumptively identify the streptococci. Among the beta-hemolytic streptococci; 98% of 50 group A, 98% of 46 group B, and 100% of 70 strains that were not group A, B, or D were correctly identified by the new presumptive test scheme. Among the non-beta-hemolytic streptococci; 96% of 74 group D enterococcal, 100% of 30 group D nonenterococcal, and 82% of 112 viridans strains were correctly identified by the new presumptive test scheme.

A Rapid Staphylococcal Coagglutination Technic to Differentiate Group A from Other Streptococcal Groups

A rapid staphylococcal coagglutination technic for differentiating group A from other streptococcal groups properly classified 500 isolates of beta-hemolytic streptococci when compared with the results of a reference capillary precipitin test. Presumptive classification of the same 500 isolates by bacitracin susceptibility produced a significant number of incorrect identifications. The findings further document the inherent inaccuracy of the bacitracin susceptibility test and suggest that the staphylococcal coagglutination technic may be a simple, accurate, inexpensive alternative for identification of group A streptococci.

Bacitracin differentiation for presumptive identification of group A β-hemolytic streptococci: Comparison of primary and purified plate testing

The detection of group A beta-hemolytic streptococci by bacitracin susceptibility was examined, with the presumptive identification on primary blood agar plates compared with that on pure subcultured plates. A total of 638 group A streptococcus isolates was recovered from 4,632 throat swab specimens. Only about half of these isolates were identifiable on the primary plate, compared with 96% identifiable on pure subcultured plates. The unreliable results with bacitracin disk testing on primary plates were due to (1) too few colonies present in the vicinity of the disk or (2) colonies overgrown by oropharyngeal flora.

Presumptive identification of group A, B, and D streptococci.

A battery of five tests was used for presumptive identification of the pathogenic streptococci. The non-serological methods included determination of hemolysis for all strains, bacitracin susceptibility for group A streptococci, hippurate hydrolysis by group B streptococci, and bile-esculin reaction for group D streptococci. Enterococcal group D streptococci were differentiated from non-enterococcal group D streptococci by 6.5% NaCl tolerance. Two other categories of streptococci resulted: beta-hemolytic streptococci non-groups A, B, or D; and alpha- or nonhemolytic streptococci, not enterococci, not further identified (viridans streptococci). The tests were used as a battery and not as single entities. In this manner more than 99% of the group A, 99% of the group B, 81% of the beta-hemolytic streptococci non-group A, B, or D, 99% of the group D enterococci, 97% of the group D non-enterococci, and 94% of the viridans streptococci were correctly identified.