Draft Genome Sequences of Viridans Streptococci Causing Bacterial Endophthalmitis in Humans

The viridans streptococci are a group of bacteria that are commensals of the oral cavity and pharynx. These species tend to cause severe cases of bacterial endophthalmitis with poor prognoses but remain largely uncharacterized in this context. Here, we report the whole-genome sequences of 21 strains of viridans streptococci isolated from endophthalmitis in humans.

Impact of frailty on elderly patients with infective endocarditis

Introduction Frailty studies focused on patients with infective endocarditis (IE) are scarce and its potential impact on patient outcomes is not well known. The aim of this study is to describe the clinical profile and prognosis of elderly patients with IE, comparing patients who met the frailty criteria versus those who did not. Methods A total of 121 cases of confirmed IE were consecutively collected in three tertiary hospitals between 2017 and 2019. The patients were classified into two groups: Group I (n=49), patients with IE who met the Frail criteria for frailty, and Group II (n=72), those patients without frailty by this scale. Results The median age of our cohort was 77 years (69–82), and 62.8% were men. Frail patients were older than those in Group II, as shown in Table 1. Regarding comorbidity, chronic anemia (40.8% vs 25%; p<0.060) was more common in Group I, as well as rheumatic manifestations at admission (12.2% vs 1.4%; p=0.014). The most frequently isolated microorganisms were S. aureus (n=25), coagulase negative staphylococci (n=25), viridans group streptococci (n=14), and enterococci (n=14). Enterococci (16.3% vs 8.3%, p=0.177) and non-viridans streptococci (10.2% vs 2.8%); p=0.086) were more frequent in frail patients. Vegetation (79.6% vs 80.6%; p=0.896) and periannular complications (24.5% vs 29.2%; p=0.571) were similar in both groups. No significant differences were found regarding the location of the infection. The incidence of in-hospital complications was similar between both groups. Frail patients underwent surgery less frequently than those in Group II, and had higher predicted mortality on surgical risk scale scores. However, the percentage of patients who met the surgical criteria and were considered inoperable was similar (33.3% vs 26.2%; p=0.415). In-hospital mortality was similar in both groups. When analyzing in-hospital mortality according to the therapeutic strategy in Group I, a mortality of 34.5% was observed in frail patients with conservative medical treatment, compared to 47.1% in those patients who underwent surgery in the same group. One third of our patients received outpatient antibiotic treatment, being significantly more frequent in Group I (39.6% vs 29.0%; p=0.232). Conclusions The elderly patients with IE and frailty criteria were older and more frequently had rheumatic symptoms at admission. Enterococci and non-viridans streptococci were isolated more frequently than in non-frailty patients. Surgery was less performed among frail patients, who had a higher predicted surgical risk. Although complications and in-hospital mortality were similar between both groups, in the group of frail patients, those with conservative management showed lower mortality compared to surgery. FUNDunding Acknowledgement Type of funding sources: None.

The Pathogenic Bacteria of Deep Neck Infection in Patients with Type 1 Diabetes, Type 2 Diabetes, and Without Diabetes from Chang Gung Research Database

Deep neck infection (DNI) is a lethal emergent condition. Patients with types 1 and 2 diabetes mellitus (T1DM and T2DM, respectively) are predisposed to DNI and have poorer prognoses. The mainstay of the treatment is surgical drainage and antibiotics; however, the pathogenic bacteria of T1DM-DNI have not been studied before. We obtained the data of 8237 patients with DNI who were hospitalized from 2004 to 2015 from the Chang Gung Research Database, which contains multi-institutional medical records in Taiwan. Using diagnostic codes, we classified them into T1DM-DNI, T2DM-DNI, and non-DM-DNI and analyzed their pathogenic bacteria, disease severity, treatment, and prognosis. The top three facultative anaerobic or aerobic bacteria of T1DM-DNI were Klebsiella pneumoniae (KP, 40.0%), Viridans Streptococci (VS, 22.2%), and methicillin-sensitive Staphylococcus aureus (MSSA, 8.9%), similar for T2DM (KP, 32.2%; VS, 23.3%; MSSA, 9.5%). For non-DM-DNI, it was VS (34.6%), KP (9.8%), and coagulase-negative Staphylococci (8.7%). The order of anaerobes for the three groups was Peptostreptococcus micros, Prevotella intermedia, and Peptostreptococcus anaerobius. Patients with T1DM-DNI and T2DM-DNI had higher white blood cell (WBC) counts and C-reactive protein (CRP) levels, more cases of surgery, more cases of tracheostomy, longer hospital stays, more mediastinal complications, and higher mortality rates than those without DM-DNI. Patients in the death subgroup in T1DM-DNI had higher WBC counts, band forms, and CRP levels than those in the survival subgroup. Patients with DM-DNI had more severe disease and higher mortality rate than those without DM-DNI. KP and Peptostreptococcus micros are the leading pathogens for both patients with T1DM-DNI and those with T2DM-DNI. Clinicians should beware of high serum levels of infection markers, which indicate potential mortality.

In vitro evaluation of disease-modifying antirheumatic drugs against rheumatoid arthritis associated pathogens of the oral microflora

ObjectivesIn the past, the human microbiome has consistently been associated with rheumatoid arthritis (RA) and disease activity. Here, we investigate the antimicrobial activity of disease-modifying antirheumatic drugs (DMARDs) against typical representatives of the oral microflora that have been associated with RA.MethodsDMARDs were screened for antimicrobial activity against bacteria that are associated with the pathogenesis of the disease and/or frequently isolated from the oral microflora of patients with RA. Screening was done by an agar diffusion assay and minimum inhibitory concentrations (MICs) of antimicrobial active substances were then determined by broth dilution.ResultsAurothiomalate and sulfasalazine demonstrated broad-spectrum antimicrobial activity, but with MICs ranging from 18 to >280 µg/mL and 150 to >600 µg/mL, respectively, only at supratherapeutic concentrations. Methotrexate showed antimicrobial activity only against Fusobacterium nucleatum and Viridans streptococci. The corresponding MICs were 3.75 to >30 µg/mL and 0.5–15 µg/mL, respectively, thus at least for streptococci, within the therapeutically achievable range. No other DMARD tested showed antimicrobial activity in the agar diffusion screening assay.ConclusionMethotrexate, sulfasalazine and aurothiomalate showed antimicrobial activity against a broad spectrum of RA associated pathogens of the oral microflora. While methotrexate showed relevant antimicrobial activity, and to a more limited extent aurothiomalate, sulfasalazine was active only at far supratherapeutic systemic concentrations. Nevertheless, given the highly species-dependent antimicrobial activity and the multiple ways it can affect the human microbiome, our results suggest a link between antimicrobially active antirheumatic drugs and their potential effect in the treatment of RA.

Structural and functional analysis of the C-terminal region of Streptococcus gordonii SspB

Streptococcus gordonii is a member of the viridans streptococci and is an early colonizer of the tooth surface. Adherence to the tooth surface is enabled by proteins present on the S. gordonii cell surface, among which SspB belongs to one of the most well studied cell-wall-anchored adhesin families: the antigen I/II (AgI/II) family. The C-terminal region of SspB consists of three tandemly connected individual domains that display the DEv-IgG fold. These C-terminal domains contain a conserved Ca2+-binding site and isopeptide bonds, and they adhere to glycoprotein 340 (Gp340; also known as salivary agglutinin, SAG). Here, the structural and functional characterization of the C123 SspB domain at 2.7 Å resolution is reported. Although the individual C-terminal domains of Streptococcus mutans AgI/II and S. gordonii SspB show a high degree of both sequence and structural homology, superposition of these structures highlights substantial differences in their electrostatic surface plots, and this can be attributed to the relative orientation of the individual domains (C1, C2 and C3) with respect to each other and could reflect their specificity in binding to extracellular matrix molecules. Studies further confirmed that affinity for Gp340 or its scavenger receptor cysteine-rich (SRCR) domains requires two of the three domains of C123 SspB, namely C12 or C23, which is different from AgI/II. Using protein–protein docking studies, models for this observed functional difference between C123 SspB and C123 AgI/II in their binding to SRCR1 are presented.

A New Species-Specific Typing Method for Salivarius Group Streptococci Based on the Dephospho-Coenzyme A Kinase (coaE) Gene Sequencing

Viridans streptococci are a group of α-hemolytic streptococcal species. They are mainly commensals, most abundant in the mouth supporting oral health. But they also include important human pathogens such as Streptococcus pneumoniae. Identification and molecular typing of viridans group streptococci are challenging, especially for members of the salivarius group. In this study, we developed a single-locus molecular typing method that is able to differentiate among the highly phylogenetically related members of the salivarius group (S. salivarius, S. vestibularis and S. thermophilus) and might support differentiation in other groups as well. This typing approach is based on the amplification and sequence analysis of the housekeeping gene dephospho-coenzyme A kinase (coaE), a gene with unrecognized taxonomic potential to date. Here, we analysed coaE gene sequences of 154 publicly available genomes and of 30 salivarius group isolates of our own collection that together belong to 20 different gram-positive bacterial (sub) species. Our results revealed that the coaE phylogeny distinguished between streptococcal and non-streptococcal genomes and that coaE gene sequences were species-specific. In contrast to MALDI-TOF MS performance, the coaE typing was able to precisely identify the phylogenetically very closely related members of the salivarius group.

Development of high level daptomycin resistance (HLDR) in Abiotrophia and Granulicatella spp isolates from patients with infective endocarditis (IE).

Abiotrophia and Granulicatella species are fastidious organisms, representing around 1%–3% of infective endocarditis (IE). Little is known about the optimal antibiotic treatment of these species, and daptomycin has been suggested as a therapeutic option. We describe the antimicrobial profile in Abiotrophia and Granulicatella IE isolates, investigate high-level daptomycin resistance (HLDR) development and evaluate daptomycin activity in combination therapy. In vitro studies with 16 IE strains (6 A. defectiva , 9 G. adiacens and 1 G. elegans ) were performed using microdilution to determine minimal inhibitory concentration (MIC) and time-kill methodology to evaluate combination therapy. Daptomycin non-susceptibility (DNS; MIC≥ 2 mg/L) and HLDR (MIC≥256 mg/L) were based on existing Clinical and Laboratory Standards (CLSI) breakpoints for viridans streptococci. All isolates were susceptible to vancomycin: G. adiacens was more susceptible to penicillin and ampicillin than A. defectiva (22% vs. 0%, and 67% vs. 33%) but less susceptible to ceftriaxone and daptomycin (56% vs. 83%, and 11% vs. 50%). HLDR developed in both A. defectiva (33%) and G. adiacens (78%) after 24h exposure to daptomycin. Combination therapy did not prevent the development of daptomycin resistance with ampicillin (2/3 strains), gentamicin (2/3 strains), ceftriaxone (2/3 strains) or ceftaroline (2/3 strains). Once developed, HLDR was stable for a prolonged time (>3 weeks) in G. adiacens , whereas in A. defectiva the HLDR it reversed to baseline MIC at day 10. This study is first to demonstrate rapid HLDR development in Abiotrophia and Granulicatella species in vitro . Resistance was stable, and most combination therapies did not prevent it.

Commensal Streptococcus mitis produces two different lipoteichoic acids of type I and type IV

The opportunistic pathogen Streptococcus mitis possesses, like other members of the Mitis group of viridans streptococci, phosphorylcholine (P-Cho)-containing teichoic acids (TAs) in its cell wall. Bioinformatic analyses predicted the presence of TAs that are almost identical with those identified in the pathogen S. pneumoniae, but a detailed analysis of S. mitis lipoteichoic acid (LTA) was not performed to date. Here we determined the structures of LTA from two S. mitis strains, the high-level beta-lactam and multiple antibiotic resistant strain B6 and the penicillin-sensitive strain NCTC10712. In agreement with bioinformatic predictions we found that the structure of one LTA (type IV) was like pneumococcal LTA, except the exchange of a glucose moiety with a galactose within the repeating units. Further genome comparisons suggested that the majority of S. mitis strains should contain the same type IV LTA as S. pneumoniae, providing a more complete understanding of the biosynthesis of these P-Cho-containing TAs in members of the Mitis group of streptococci. Remarkably, we observed besides type IV LTA an additional polymer belonging to LTA type I in both investigated S. mitis strains. This LTA consists of β-galactofuranosyl-(1,3)-diacylglycerol as glycolipid anchor and a poly-glycerol-phosphate chain at the O-6 position of the furanosidic galactose. Hence, these bacteria are capable of synthesizing two different LTA polymers, most likely produced by distinct biosynthesis pathways. Our bioinformatics analysis revealed the prevalence of the LTA synthase LtaS, most probably responsible for the second LTA version (type I), amongst S. mitis and S. pseudopneumoniae strains.

Direct DNA Sequencing-Based Analysis of Microbiota Associated with Hematological Malignancies in the Eastern Province of Saudi Arabia

Introduction. Bloodstream infections (BSI) among patients with hematological malignancies (HM) could predispose them to higher morbidity and mortality for various underlying conditions. Several microorganisms, either pathogenic or opportunistic normal human flora, could cause severe bacteremia and septicemia. While conventional methods have their own limitations, molecular methods such as next-generation sequencing (NGS) can detect these blood infections with more reliability, specificity, and sensitivity, in addition to information on microbial population landscape. Methodology. Blood samples from HM patients ( n = 50 ) and volunteer blood donor control individuals with no HM ( n = 50 ) were subjected to 16S rRNA gene amplification using standard PCR protocols. A metagenomic library was prepared, and NGS was run on a MiSeq (Illumina) sequencer. Sequence reads were analyzed using MiSeq Reporter, and microbial taxa were aligned using the Green Genes library. Results. 82% of the patients showed BSI with Gram-negative bacteria as the most predominant group. E. coli comprised a major chunk of the bacterial population (19.51%), followed by K. pneumoniae (17.07%). The CoNS and Viridans Streptococci groups are 17.07% and 14.63%, respectively. Other major species were S. aureus (9.75%), P. aeruginosa (7.31%), A. baumannii (4.87%), E. cloacae (4.87%), and P. mirabilis (4.87%). 34.14% of the cases among patients showed a Gram-positive infection, while 14.63% showed polymicrobial infections. Conclusion. Most of the BSI in patients were characterized by polymicrobial infections, unlike the control samples. Molecular methods like NGS showed robust, fast, and specific identification of infectious agents in BSI in HM, indicating the possibility of its application in routine follow-up of such patients for infections.