This report describes mutations in genes responsible for cell deformities in haemophili under beta-lactam pressure in vitro. Light and transmission electron microscopy confirmed a hypothesis regarding changes in the shape of haemophili that had become more filamentous in the presence of ampicillin (2 mg/L) and cefuroxime (8 mg/L) after 30 days of serial passage. Short-axis size increased by 28% (from 0.767 to 1.06 µm) and long-axis length increased by 54% (from 1 to 2.175 µm). Additionally, whole-genome sequencing analysis (Illumina platform, software PROKKA) revealed a variety of mutations in genes responsible for cell morphology in isolates examined in this study: ftsI (A1576 → C; G1154 → C; T986 → C; G1684 → C), mreB (C476 → T), mreC (A5 → G), mrdA (A1148 → G; C179 → T; G1613 → T), mrdB (T668 → G), mltC (C1016 → T) and rodA (T668 → G). The results of this study indicate that shifts in bacterial shape could play a role in the adaptation of haemophili to a new niche created by beta-lactams as a strategy of antibiotic therapy survival.
- Klíčová slova
- Beta-lactams, Haemophili, Whole-genome sequencing,
- MeSH
- ampicilin farmakologie MeSH
- antibakteriální látky * farmakologie MeSH
- beta-laktamy * farmakologie MeSH
- cefuroxim farmakologie MeSH
- genom bakteriální * MeSH
- Haemophilus influenzae * genetika účinky léků MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- mutace MeSH
- sekvenování celého genomu * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- ampicilin MeSH
- antibakteriální látky * MeSH
- beta-laktamy * MeSH
- cefuroxim MeSH
Secretory (S) IgA antibodies against severe acute respiratory syndrome (SARS)-CoV-2 are induced in saliva and upper respiratory tract (URT) secretions by natural infection and may be critical in determining the outcome of initial infection. Secretory IgA1 (SIgA1) is the predominant isotype of antibodies in these secretions. Neutralization of SARS-CoV-2 is most effectively accomplished by polymeric antibodies such as SIgA. We hypothesize that cleavage of SIgA1 antibodies against SARS-CoV-2 by unique bacterial IgA1 proteases to univalent Fabα antibody fragments with diminished virus neutralizing activity would facilitate the descent of the virus into the lungs to cause serious disease and also enhance its airborne transmission to others. Recent studies of the nasopharyngeal microbiota of patients with SARS-CoV-2 infection have revealed significant increases in the proportions of IgA1 protease-producing bacteria in comparison with healthy subjects. Similar considerations might apply also to other respiratory viral infections including influenza, possibly explaining the original attribution of influenza to Haemophilus influenzae, which produces IgA1 protease.
- Klíčová slova
- COVID-19, SARS-CoV-2, bacterial IgA1 protease, immunoglobulin A, mucosal immunity,
- MeSH
- Bacteria enzymologie MeSH
- COVID-19 * přenos imunologie virologie mikrobiologie MeSH
- Haemophilus influenzae enzymologie imunologie MeSH
- imunoglobulin A sekreční * metabolismus MeSH
- imunoglobulin A imunologie MeSH
- lidé MeSH
- nazofarynx mikrobiologie virologie MeSH
- neutralizující protilátky imunologie MeSH
- protilátky virové imunologie MeSH
- SARS-CoV-2 * imunologie MeSH
- serinové endopeptidasy metabolismus imunologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- IgA-specific serine endopeptidase MeSH Prohlížeč
- imunoglobulin A sekreční * MeSH
- imunoglobulin A MeSH
- neutralizující protilátky MeSH
- protilátky virové MeSH
- serinové endopeptidasy MeSH
BACKGROUND: Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. One of the mechanisms of resistance to β-lactams is the alteration of the transpeptidase region of penicillin binding protein 3 (PBP3) which is caused by mutations in the ftsI gene. It was shown that exposure to beta-lactams has a stimulating effect on increase of prevalence of H. influenzae strains with the non-enzymatic mechanism of resistance. OBJECTIVES: The aim of our study was to compare the mutational potential of ampicillin and cefuroxime in H. influenzae strains, determination of minimum inhibitory concentration and the evolution of mutations over time, focusing on amino acid substitutions in PBP3. METHODS: 30 days of serial passaging of strains in liquid broth containing increasing concentrations of ampicillin or cefuroxime was followed by whole-genome sequencing. RESULTS: On average, cefuroxime increased the minimum inhibitory concentration more than ampicillin. The minimum inhibitory concentration was increased by a maximum of 32 fold. Substitutions in the PBP3 started to appear after 15 days of passaging. In PBP3, cefuroxime caused different substitutions than ampicillin. CONCLUSIONS: Our experiment observed differences in mutation selection by ampicillin and cefuroxime. Selection pressure of antibiotics in vitro generated substitutions that do not occur in clinical strains in the Czech Republic.
- Klíčová slova
- Ampicillin, Cefuroxime, Haemophilus, PBP3, WGS,
- MeSH
- ampicilin * farmakologie MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny genetika metabolismus MeSH
- cefuroxim * farmakologie MeSH
- Haemophilus influenzae * genetika účinky léků MeSH
- hemofilové infekce mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- molekulární evoluce MeSH
- mutace * MeSH
- proteiny vázající penicilin * genetika metabolismus MeSH
- sekvenování celého genomu MeSH
- selekce (genetika) MeSH
- sériové pasážování MeSH
- substituce aminokyselin * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- ampicilin * MeSH
- antibakteriální látky * MeSH
- bakteriální proteiny MeSH
- cefuroxim * MeSH
- proteiny vázající penicilin * MeSH
BACKGROUND: The Invasive Respiratory Infection Surveillance (IRIS) Consortium was established to assess the impact of the COVID-19 pandemic on invasive diseases caused by Streptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidis, and Streptococcus agalactiae. We aimed to analyse the incidence and distribution of these diseases during the first 2 years of the COVID-19 pandemic compared to the 2 years preceding the pandemic. METHODS: For this prospective analysis, laboratories in 30 countries and territories representing five continents submitted surveillance data from Jan 1, 2018, to Jan 2, 2022, to private projects within databases in PubMLST. The impact of COVID-19 containment measures on the overall number of cases was analysed, and changes in disease distributions by patient age and serotype or group were examined. Interrupted time-series analyses were done to quantify the impact of pandemic response measures and their relaxation on disease rates, and autoregressive integrated moving average models were used to estimate effect sizes and forecast counterfactual trends by hemisphere. FINDINGS: Overall, 116 841 cases were analysed: 76 481 in 2018-19, before the pandemic, and 40 360 in 2020-21, during the pandemic. During the pandemic there was a significant reduction in the risk of disease caused by S pneumoniae (risk ratio 0·47; 95% CI 0·40-0·55), H influenzae (0·51; 0·40-0·66) and N meningitidis (0·26; 0·21-0·31), while no significant changes were observed for S agalactiae (1·02; 0·75-1·40), which is not transmitted via the respiratory route. No major changes in the distribution of cases were observed when stratified by patient age or serotype or group. An estimated 36 289 (95% prediction interval 17 145-55 434) cases of invasive bacterial disease were averted during the first 2 years of the pandemic among IRIS-participating countries and territories. INTERPRETATION: COVID-19 containment measures were associated with a sustained decrease in the incidence of invasive disease caused by S pneumoniae, H influenzae, and N meningitidis during the first 2 years of the pandemic, but cases began to increase in some countries towards the end of 2021 as pandemic restrictions were lifted. These IRIS data provide a better understanding of microbial transmission, will inform vaccine development and implementation, and can contribute to health-care service planning and provision of policies. FUNDING: Wellcome Trust, NIHR Oxford Biomedical Research Centre, Spanish Ministry of Science and Innovation, Korea Disease Control and Prevention Agency, Torsten Söderberg Foundation, Stockholm County Council, Swedish Research Council, German Federal Ministry of Health, Robert Koch Institute, Pfizer, Merck, and the Greek National Public Health Organization.
AIM: To assess the trends and changes in the incidence of invasive disease caused by Haemophilus influenzae in the Czech Republic (CR) between 1999 and 2020 with regard to the introduction of childhood vaccination against H. influenzae serotype b (Hib) in 2001. Characterization of strains by multilocus sequence typing (MLST) and search for correlations between serotypes, sequence types, and patient groups or clinical manifestations of the disease. MATERIAL AND METHODS: A total of 623 invasive H. influenzae strains from surveillance of invasive Haemophilus disease in the Czech Republic were analysed. All strains were biotyped based on phenotypic characteristics and serotyped using slide agglutination with specific a-f antisera. Three hundred and eighty-three strains from the collection of the National Reference Laboratory for Haemophilus Infections (NRL HEM) originating from surveillance in the CR were analysed by MLST and assigned to sequence types (ST). For analyses, the dataset was supplemented with five strains from the PubMLST database of serotypes rarely or not at all found in the CR. Similarity calculations based on MLST and strain (serotype, biotype, ST) and patient (diagnosis, sex, age) data were performed in BioNumerics 7.6. RESULTS: After the introduction of Hib vaccination in 2001, a dramatic decline of more than 90% was observed in invasive Hib disease over the following years. Between 1999 and 2020, a total of 623 cases of invasive disease caused by H. influenzae were recorded in the CR, with about 20 cases reported annually in recent years. At present, the dominant agents causing Haemophilus invasive disease in the CR are non-enveloped strains (HiNT) followed by strains of Hif and Hie serotypes. The most common manifestation of Haemophilus invasive disease in the pre-vaccination era was meningitis, while now it is sepsis. Sequence types of 383 strains from the NRL HEM collection originating from surveillance in the CR were analysed. The results showed high clonality of the encapsulated strains and diversity of HiNT strains, which is consistent with the results of others. Strain similarity analysis showed no demonstrable relationships between patient age or clinical manifestation and serotype and ST. CONCLUSION: In invasive Haemophilus disease, there has been a dramatic change as a result of Hib vaccination after 2001, with a reduction of cases caused by Hib from tens to units annually. In the last decade, the situation in the CR has been stable with no significant changes in the number of cases or in the representation of causative serotypes and is in line with the reports from other EU countries. In order to monitor further developments, it is desirable that the NRL HEM should continue the surveillance of invasive disease caused by H. influenzae, including molecular biological characteristics of strains. MLST allows the characterisation of strains based on allelic variants of selected housekeeping genes, but it does not allow the association of specific H. influenzae sequence types with patient age, sex or clinical manifestations. In the future, whole genome sequencing could be a useful tool for determining the correlation between the disease and specific strains.
- Klíčová slova
- Epidemiology, Haemophilus influenzae, MLST, biotyping, invasive disease, serotyping, surveillance, type 1 diabetes,
- MeSH
- Haemophilus influenzae * genetika MeSH
- hemofilové infekce * epidemiologie MeSH
- kojenec MeSH
- lidé MeSH
- multilokusová sekvenční typizace MeSH
- sérotypizace MeSH
- Check Tag
- kojenec MeSH
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika epidemiologie MeSH
While the inhalation of Thymus vulgaris L. essential oil (EO) is commonly approved for the treatment of mild respiratory infections, there is still a lack of data regarding the antimicrobial activity and chemical composition of its vapours. The antibacterial activity of the three T. vulgaris EOs against respiratory pathogens, including Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pyogenes, was assessed in both liquid and vapour phases using the broth microdilution volatilisation (BMV) method. With the aim of optimising a protocol for the characterisation of EO vapours, their chemical profiles were determined using two headspace sampling techniques coupled with GC/MS: solid-phase microextraction (HS-SPME) and syringe headspace sampling technique (HS-GTS). All EO sample vapours exhibited antibacterial activity with minimum inhibitory concentrations (MIC) ranging from 512 to 1024 μg/mL. According to the sampling technique used, results showed a different distribution of volatile compounds. Notably, thymol was found in lower amounts in the headspace-peak percentage areas below 5.27% (HS-SPME) and 0.60% (HS-GTS)-than in EOs (max. 48.65%), suggesting that its antimicrobial effect is higher in vapour. Furthermore, both headspace sampling techniques were proved to be complementary for the analysis of EO vapours, whereas HS-SPME yielded more accurate qualitative results and HS-GTS proved a better technique for quantitative analysis.
- Klíčová slova
- antimicrobial activity, broth microdilution, headspace analysis, respiratory infections, thyme, vapour phase,
- MeSH
- antibakteriální látky chemie izolace a purifikace farmakologie MeSH
- Haemophilus influenzae účinky léků MeSH
- mikrobiální testy citlivosti MeSH
- mikroextrakce na pevné fázi * MeSH
- oleje prchavé chemie izolace a purifikace farmakologie MeSH
- plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
- Staphylococcus aureus účinky léků MeSH
- Streptococcus pyogenes účinky léků MeSH
- Thymus (rostlina) chemie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antibakteriální látky MeSH
- oleje prchavé MeSH
In this study, a new broth macrodilution volatilization method for the simple and rapid determination of the antibacterial effect of volatile agents simultaneously in the liquid and vapor phase was designed with the aim to assess their therapeutic potential for the development of new inhalation preparations. The antibacterial activity of plant volatiles (β-thujaplicin, thymohydroquinone, thymoquinone) was evaluated against bacteria associated with respiratory infections (Haemophilus influenzae, Staphylococcus aureus, Streptococcus pneumoniae, Streptococcus pyogenes) and their cytotoxicity was determined using a modified thiazolyl blue tetrazolium bromide assay against normal lung fibroblasts. Thymohydroquinone and thymoquinone possessed the highest antibacterial activity against H. influenzae, with minimum inhibitory concentrations of 4 and 8 µg/mL in the liquid and vapor phases, respectively. Although all compounds exhibited cytotoxic effects on lung cells, therapeutic indices (TIs) suggested their potential use in the treatment of respiratory infections, which was especially evident for thymohydroquinone (TI > 34.13). The results demonstrate the applicability of the broth macrodilution volatilization assay, which combines the principles of broth microdilution volatilization and standard broth macrodilution methods. This assay enables rapid, simple, cost- and labor-effective screening of volatile compounds and overcomes the limitations of assays currently used for screening of antimicrobial activity in the vapor phase.
- Klíčová slova
- antimicrobial, cytotoxicity, macrodilution method, respiratory infections, thymohydroquinone, thymoquinone, vapor phase, volatile compound, β-thujaplicin,
- MeSH
- antibakteriální látky analýza chemie farmakologie MeSH
- aplikace inhalační MeSH
- Bacteria účinky léků MeSH
- benzochinony aplikace a dávkování farmakologie MeSH
- Haemophilus influenzae účinky léků MeSH
- mikrobiální testy citlivosti MeSH
- monoterpeny aplikace a dávkování farmakologie MeSH
- oleje prchavé chemie farmakologie MeSH
- Staphylococcus aureus účinky léků MeSH
- Streptococcus pneumoniae účinky léků MeSH
- Streptococcus pyogenes účinky léků MeSH
- těkavé organické sloučeniny chemie farmakologie MeSH
- thymol aplikace a dávkování analogy a deriváty farmakologie MeSH
- tropolon aplikace a dávkování analogy a deriváty farmakologie MeSH
- volatilizace MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antibakteriální látky MeSH
- benzochinony MeSH
- beta-thujaplicin MeSH Prohlížeč
- monoterpeny MeSH
- oleje prchavé MeSH
- těkavé organické sloučeniny MeSH
- thymohydroquinone MeSH Prohlížeč
- thymol MeSH
- thymoquinone MeSH Prohlížeč
- tropolon MeSH
BACKGROUND: Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis, which are typically transmitted via respiratory droplets, are leading causes of invasive diseases, including bacteraemic pneumonia and meningitis, and of secondary infections subsequent to post-viral respiratory disease. The aim of this study was to investigate the incidence of invasive disease due to these pathogens during the early months of the COVID-19 pandemic. METHODS: In this prospective analysis of surveillance data, laboratories in 26 countries and territories across six continents submitted data on cases of invasive disease due to S pneumoniae, H influenzae, and N meningitidis from Jan 1, 2018, to May, 31, 2020, as part of the Invasive Respiratory Infection Surveillance (IRIS) Initiative. Numbers of weekly cases in 2020 were compared with corresponding data for 2018 and 2019. Data for invasive disease due to Streptococcus agalactiae, a non-respiratory pathogen, were collected from nine laboratories for comparison. The stringency of COVID-19 containment measures was quantified using the Oxford COVID-19 Government Response Tracker. Changes in population movements were assessed using Google COVID-19 Community Mobility Reports. Interrupted time-series modelling quantified changes in the incidence of invasive disease due to S pneumoniae, H influenzae, and N meningitidis in 2020 relative to when containment measures were imposed. FINDINGS: 27 laboratories from 26 countries and territories submitted data to the IRIS Initiative for S pneumoniae (62 837 total cases), 24 laboratories from 24 countries submitted data for H influenzae (7796 total cases), and 21 laboratories from 21 countries submitted data for N meningitidis (5877 total cases). All countries and territories had experienced a significant and sustained reduction in invasive diseases due to S pneumoniae, H influenzae, and N meningitidis in early 2020 (Jan 1 to May 31, 2020), coinciding with the introduction of COVID-19 containment measures in each country. By contrast, no significant changes in the incidence of invasive S agalactiae infections were observed. Similar trends were observed across most countries and territories despite differing stringency in COVID-19 control policies. The incidence of reported S pneumoniae infections decreased by 68% at 4 weeks (incidence rate ratio 0·32 [95% CI 0·27-0·37]) and 82% at 8 weeks (0·18 [0·14-0·23]) following the week in which significant changes in population movements were recorded. INTERPRETATION: The introduction of COVID-19 containment policies and public information campaigns likely reduced transmission of S pneumoniae, H influenzae, and N meningitidis, leading to a significant reduction in life-threatening invasive diseases in many countries worldwide. FUNDING: Wellcome Trust (UK), Robert Koch Institute (Germany), Federal Ministry of Health (Germany), Pfizer, Merck, Health Protection Surveillance Centre (Ireland), SpID-Net project (Ireland), European Centre for Disease Prevention and Control (European Union), Horizon 2020 (European Commission), Ministry of Health (Poland), National Programme of Antibiotic Protection (Poland), Ministry of Science and Higher Education (Poland), Agencia de Salut Pública de Catalunya (Spain), Sant Joan de Deu Foundation (Spain), Knut and Alice Wallenberg Foundation (Sweden), Swedish Research Council (Sweden), Region Stockholm (Sweden), Federal Office of Public Health of Switzerland (Switzerland), and French Public Health Agency (France).
- MeSH
- analýza přerušované časové série MeSH
- bakteriální infekce epidemiologie přenos MeSH
- COVID-19 * prevence a kontrola MeSH
- Haemophilus influenzae MeSH
- incidence MeSH
- infekce dýchací soustavy epidemiologie MeSH
- lidé MeSH
- Neisseria meningitidis MeSH
- prospektivní studie MeSH
- Streptococcus agalactiae MeSH
- Streptococcus pneumoniae MeSH
- surveillance populace MeSH
- veřejné zdravotnictví - praxe MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Haemophilus Influenza leads to respiratory infections such as sinusitis, acute otitis media, pneumonia and bronchitis. In addition, it causes invasive infections such as cellulite, septic arthritis, and meningitis. Therefore, quick and sensitive detection of H. influenza is of great importance in medical microbiology. In this study, a novel DNA-based bioassay was developed to the monitoring of Haemophilus influenza genome in human plasma samples using binding of pDNA with cDNA. DNA hybridization strategy was used to investigation of DNAs binding. For this purpose, silver nanoparticle doped graphene quantum dots inks functionalized by D-penicillamine (Ag NPs-DPA-GQDs) were synthesized and deposited on the surface of glass carbon electrode (GCE). Also, gold nanoparticles functionalized with cysteamine (CysA-AuNPs) were deposited on the surface of the Ag-DPA-GQDs modified GCE. Afterward, thiolated DNA probe was immobilized on the surface of the modified electrode. DNA hybridization was monitored using square wave voltammetry (SWV) technique. Engineered genosensor indicated good performance with high specificity and sensitivity for detection of Haemophilus influenza genome. Under optimal conditions, linear range and low limit of quantitation (LLOQ) were obtained as target concentrations ranging from 1 pM-1 ZM and 1 ZM, respectively. The designed biosensor also showed high capability of discriminating one-base, two-base and three-base mismatched sequences. Also, the prepared genosensor could be easily regenerated and reused to evaluate hybridization process.
- Klíčová slova
- Bioassay, DNA hybridization, Haemophilus influenza, Nucleic acid binding, Silver nanoparticles,
- MeSH
- biosenzitivní techniky MeSH
- elektrochemické techniky MeSH
- genom bakteriální * MeSH
- Haemophilus influenzae genetika MeSH
- hybridizace nukleových kyselin metody MeSH
- komplementární DNA * MeSH
- kovové nanočástice ultrastruktura MeSH
- kvantové tečky MeSH
- lidé MeSH
- reprodukovatelnost výsledků MeSH
- volné cirkulující nukleové kyseliny * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- komplementární DNA * MeSH
- volné cirkulující nukleové kyseliny * MeSH
A sensitive and specific approach was developed for the determination of Haemophilus influenza using DNA based bio-assay. In this study, citrate capped silver nanoparticle was synthesized and employed for bioconjugation with pDNA toward target sequences detection. In this study, synthesized probe (SH-5'-AAT TTT CCA ACT TTT TCA CCT GCA T-3') of Haemophilus influenza was detected with great sensitivity and selectivity after hybridization with cDNA (5'-ATG CAG GTG AAA AAG TTG GAA AAT T-3'). Regarding to the obtained results, the low limit of quantification (LLOQ) of DNA sample was 1 ZM using 15 μL of probe and 200 μL of Cit/AgNPs. According to ultra-sensitivity of the fabricated optical DNA-based bio-assay, it has potential for bacterial determination both in clinical and environmental specimens. To evaluate the selectivity of developed DNA based biosensor, three mismatch sequences were applied. Finally, the designed genosensor is a significant diagnostic strategy for detection of Haemophilus influenza with great selectivity.
- Klíčová slova
- Citrate capped silver nanoparticle, DNA bio-assay, Fluorescence, Genosensor, Haemophilus influenza, Human biofluids,
- MeSH
- biosenzitivní techniky přístrojové vybavení metody MeSH
- biotest MeSH
- DNA bakterií analýza genetika MeSH
- DNA sondy chemie genetika MeSH
- Haemophilus influenzae genetika izolace a purifikace MeSH
- hybridizace nukleových kyselin MeSH
- kovové nanočástice chemie MeSH
- kyselina citronová chemie MeSH
- lidé MeSH
- limita detekce MeSH
- senzitivita a specificita MeSH
- stříbro chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA bakterií MeSH
- DNA sondy MeSH
- kyselina citronová MeSH
- stříbro MeSH
