Case Report: Streptococcus Suis Meningitis Diagnosed in a HIV-Infected Patient With Cryptococcal Meningitis Using Next-Generation Sequencing

Background: Streptococcus suis has been recognized as a zoonotic pathogen that may cause infections in humans. Although rarely described, it is not surprising that both cryptococcal and streptococcus suis meningitis infections can co-exist in a HIV-infected patient with a low CD4 count. However, a fast and accurate diagnose of meningitis of multipathogenic infections is challenging. In this report, we describe such a case of a HIV-infected patient with meningitis of multipathogenic infections.Case Presentation: The patient was a 34-year-old Chinese male who was diagnosed with cryptococcal meningitis and HIV at the same time about 1 year ago. During the same time period, he had received (with good compliance) fluconazole and tenofovir-lamivudine- dolutegravir based antiretroviral therapy (ART). However, symptom of progressively worsening occipital headache appeared after he was exposed to a truck which was used for transporting pigs. Initial workup indicated an increase of the cerebrospinal fluid (CSF) opening pressure (OP) and an increase in the number of lymphocytes and proteins in CSF. A magnetic resonance imaging (MRI) scan revealed that partial cerebellar surface enhancement. The cryptococcus capsular antigen test of CSF was positive. The results of the India Ink microscopy for cryptococcus, nucleic acid of CMV and EBV and mycobacterium tuberculosis (MTB) tests of CSF were negative. The results of the bacteria and fungi smear and culture of CSF were also negative. Eventually, streptococcus suis was detected using next-generation sequencing (NGS) in CSF. The diagnosis of Streptococcus suis meningitis was made based on the patient's contact history with carrier pigs and the clinical findings addressed above. The treatment of 2 weeks of intravenous ceftriaxone and 1 week of oral moxifloxacin resulted in improvement of the condition of CSF. The anti-fungal treatment using fluconazole continued until the CFS OP went down to a normal level and the cryptococcus capsular antigen test of CSF was negative 6 months later.Conclusion: This case highlights that NGS might be beneficial to HIV-infected patients who have meningitis with negative CSF culture results. Multiple etiologies for such condition in the immunocompromised patients must be taken into consideration and early stage NGS is recommended.

Optimised Heterologous Expression and Functional Analysis of the Yersinia pestis F1-Capsular Antigen Regulator Caf1R

The bacterial pathogen, Yersinia pestis, has caused three historic pandemics and continues to cause small outbreaks worldwide. During infection, Y. pestis assembles a capsule-like protective coat of thin fibres of Caf1 subunits. This F1 capsular antigen has attracted much attention due to its clinical value in plague diagnostics and anti-plague vaccine development. Expression of F1 is tightly regulated by a transcriptional activator, Caf1R, of the AraC/XylS family, proteins notoriously prone to aggregation. Here, we have optimised the recombinant expression of soluble Caf1R. Expression from the native and synthetic codon-optimised caf1R cloned in three different expression plasmids was examined in a library of E. coli host strains. The functionality of His-tagged Caf1R was demonstrated in vivo, but insolubility was a problem with overproduction. High levels of soluble MBP-Caf1R were produced from codon optimised caf1R. Transcriptional-lacZ reporter fusions defined the PM promoter and Caf1R binding site responsible for transcription of the cafMA1 operon. Use of the identified Caf1R binding caf DNA sequence in an electrophoretic mobility shift assay (EMSA) confirmed correct folding and functionality of the Caf1R DNA-binding domain in recombinant MBP-Caf1R. Availability of functional recombinant Caf1R will be a valuable tool to elucidate control of expression of F1 and Caf1R-regulated pathophysiology of Y. pestis.

Experience of Using a Complex Antigenic Preparation of the Plague Microbe to Assess the Severity of a Specific Anti-Plague Response

Background. Improving the methodology of immunological monitoring in natural foci of plague in the Russian Federation and adjacent territories to increase the effectiveness of epidemiological surveillance of plague is an urgent line of research. The lack of correlation between the production of specific antibodies to the capsular antigen (F1) ofthe plague microbe with other indicators of the state of cellular defense reactivity indicates the need to search for new informative and accessible markers for assessing anti-plague immunity.Objective: to evaluate possibility of using the complex preparation (F1 and cell membranes) evaluate the possibilities of using an artificial antigenic complex based on F1 and cell membranes (CM) of the plague microbe in antigen-specific tests in vitro in people vaccinated against plague.resu. The study involved 153 volunteers living in the territory enzootic for plague (the village of Khandagayty ofthe Ovyur kozhuun of the Tyva Republic and the village of Kosh-Agach of the Kosh-Agach district of the Altai Republic). The study included the determination of spontaneous and mitogen-induced production of cytokines (IFN-γ, IL-4, TNF-α) by blood cells, titers of specific IgG antibodies to the capsular antigen F1 of the plague microbe and concentrations ofthe main classes of immunoglobulins (IgM, IgG, IgA and IgE) in blood serum, as well as immunophenotyping of blood lymphocytes (CD3, CD4, CD8, CD16, CD19).Results. Comparative assessment of the level of cytokines (TNF-α, IFN-γ and IL-4) in spontaneous/induced F1+CM Y. pestis tests revealed a statistically significant increase in the production of cytokines TNF-α and IFN-γ in the antigeninduced tests compared with spontaneous (p < 0.01).Conclusion. Thus, the effectiveness of the use of artificial antigenic complex based on F1 and cell membranes ofthe plague microbe has been shown to assess the production of cytokines in antigen-specific cell tests in vitro, which justifies the need for further research.

Virulence-associated characteristics of carbapenem-resistant Klebsiella pneumoniae in hospital-acquired infections: results from a hospital in central China

Background Carbapenem-resistant Klebsiella pneumoniae (CRKP) have been a clinically significant pathogen worldwide, but related reports about their virulence features in hospital-acquired infections (HAI) are pretty lacking.Methods CRKP causing HAI were continuously collected in 2018 from a hospital in central China. Isolates identification and antimicrobial susceptibility test were done using VITEK-2 compact system or MALDI-TOF MS. String test, multilocus sequence typing, carbapenemase genes, virulence genes and capsular antigen genes detection were conducted to understand their phenotype and genetic background. As well as case datas were collected and compared to assess their virulence characteristics.Results A total of 62 isolates of CRKP from 62 patients with HAI were collected. 41 carbapenemase genestic-confirmed hypervirulent Klebsiella pneumoniae (CR-hvKP) and 21 carbapenem resistant non-hypervirulent Klebsiella pneumoniae (CR-NhvKP) were screened out. Most CRKP causing HAI were ST11 KPC-2 producing strains and maily causing pneumonia. Only for blaKPC-2 there was a significant difference between CR-hvKP and CR-NhvKP (p<0.001). No significant difference of the two group strains in resistance against amikacin, trimethoprim-sulfamethoxazoleare, cefepime, ceftazidime, imipenem, piperacillin-tazobactam, colistin and tigecycline were found except levofloxacin (p<0.001), and all strains showed sensitive to tigecycline and colistin. In the CR-hvKP group, IucA (64.5%) were the most commonly detected virulence gene, followed by iroN (48.4%), prmpA2 (30.6%) and prmpA (4.8%), only 1 (2.4%) capsular serotype positive strain and 2 (4.9%) hypermucoviscosity phenotype strains were detected, while no hypermucoviscosity phenotype or capsular antigen gene positive strain was detected in the CR-NhvKP group. And there was no significant difference between the two groups in age, types of infection, departmental distribution, survival time or the final outcome of infection.Conclusion ST11 KPC-2-producing Klebsiella pneumoniae are most prevalent CRKP in HAI. Virulence gene espacially iucA has a high proportion and worth paying attention to. Hypermucoviscous phenotype and virulence-associated capsular serotype in CRKP both have a low prevalence. CRKP harboring virulence genes have a higher expression of KPC-2 and less sensitive to levofloxacin than those harboring no virulence gene, and there is no significant difference for virulence manifestations between the two groups.

Adsorption of Vi Capsular Antigen of Salmonella Typhi in Chitosan–Poly (Methacrylic Acid) Nanoparticles

The development of a nanoparticulate system for the carrier antigen is now an important tool in the vaccination process, since a smaller number of doses is necessary for effective immunization. Thus, in this work a nanoparticulate system using polymers of chitosan and poly (methacrylic acid) (CS–PMAA) to adsorb the Vi antigen of Salmonella Typhi was developed. CS–PMAA nanoparticles with different proportions of chitosan and poly (methacrylic acid) were obtained and reached sizes from 123.9 ± 2.48 to 234.9 ± 2.66 nm, and spherical shapes were seen in transmission microscopy. At pH 7.2, the nanoparticles had a cationic surface charge that contributed to the adsorption of the Vi antigen. Qualitative analyses of the isolated Vi antigen were performed using Fourier-transform infrared spectroscopy, which indicated the presence of all the characteristic bands of the capsular polysaccharide, and nuclear magnetic resonance, which showed signals for the five hydrogens and the N-acetyl and O-acetyl groups which are characteristic of the Vi antigen structure. In the adsorption kinetics study, the Vi capsular antigen, contained in a phosphate buffer solution of pH 7.2, experienced 55% adsorption on the 1–1% (CS–PMAA) nanoparticles. The adsorption kinetics results showed the ability of the nanoparticulate system to adsorb the Vi antigen.