Neurological manifestations of nontuberculous mycobacteria in adults: case series and review of the literature.

Introduction: Nontuberculous mycobacteria (NTM) mediated infections are important to consider in cases with neuroinflammatory presentations. We aimed to characterize cases of NTM with neurological manifestations at the National Institutes of Health (NIH) Clinical Center and review the relevant literature. Materials and methods: Between January 1995 and December 2020, six cases were identified. Records were reviewed for demographic, clinical, and radiological characteristics. A MEDLINE search found previously reported cases. Data were extracted, followed by statistical analysis to compare two groups [cases with slow-growing mycobacteria (SGM) vs. those with rapidly growing mycobacteria (RGM)] and evaluate for predictors of survival. NIH cases were evaluated for clinical and radiological characteristics. Cases from the literature were reviewed to determine the differences between SGM and RGM cases and to identify predictors of survival. Results: Six cases from NIH were identified (age 41 ± 13, 83% male). Five cases were caused by SGM [Mycobacterium avium complex (MAC) n = 4; Mycobacterium haemophilum n = 1] and one due to RGM (Mycobacterium abscessus). Underlying immune disorders were identified only in the SGM cases [genetic (n = 2), HIV (n = 1), sarcoidosis (n = 1), and anti-interferon-gamma antibodies (n = 1)]. All cases were diagnosed using tissue analysis. A literature review found 81 reports on 125 cases (SGM n = 85, RGM n = 38, non-identified n = 2). No immune disorder was reported in 26 cases (21%). Within SGM cases, the most common underlying disease was HIV infection (n = 55, 65%), and seizures and focal lesions were more common. In RGM cases, the most common underlying condition was neurosurgical intervention or implants (55%), and headaches and meningeal signs were common. Tissue-based diagnosis was used more for SGM than RGM (39% vs. 13%, p = 0.04). Survival rates were similar in both groups (48% SGM and 55% in RGM). Factors associated with better survival were a solitary CNS lesion (OR 5.9, p = 0.01) and a diagnosis made by CSF sampling only (OR 9.9, p = 0.04). Discussion: NTM infections cause diverse neurological manifestations, with some distinctions between SGM and RGM infections. Tissue sampling may be necessary to establish the diagnosis, and an effort should be made to identify an underlying immune disorder.

A simple solid media assay for detection of synergy between bacteriophages and antibiotics.

The emergence of antibiotic-resistant bacteria (ARB) has necessitated the development of alternative therapies to deal with this global threat. Bacteriophages (viruses that target bacteria) that kill ARB are one such alternative. Although phages have been used clinically for decades with inconsistent results, a number of recent advances in phage selection, propagation, and purification have enabled a reevaluation of their utility in contemporary clinical medicine. In most phage therapy cases, phages are administered in combination with antibiotics to ensure that patients receive the standard-of-care treatment. Some phages may work cooperatively with antibiotics to eradicate ARB, as often determined using non-standardized broth assays. We sought to develop a solid media-based assay to assess cooperativity between antibiotics and phages to offer a standardized platform for such testing. We modeled the interactions that occur between antibiotics and phages on solid medium to measure additive, antagonistic, and synergistic interactions. We then tested the method using different bacterial isolates and identified a number of isolates where synergistic interactions were identified. These interactions were not dependent on the specific organism, phage family, or antibiotic used. A priori susceptibility to the antibiotic or the specific phage were not requirements to observe synergistic interactions. Our data also confirm the potential for the restoration of vancomycin to treat vancomycin-resistant Enterococcus (VRE) when used in combination with phages. Solid media assays for the detection of cooperative interactions between antibiotics and phages can be an accessible technique adopted by clinical laboratories to evaluate antibiotic and phage choices in phage therapy.IMPORTANCEBacteriophages have become an important alternative treatment for individuals with life-threatening antibiotic-resistant bacteria (ARB) infections. Because antibiotics represent the standard-of-care for treatment of ARB, antibiotics and phages often are delivered together without evidence that they work cooperatively. Testing for cooperativity can be difficult due to the equipment necessary and a lack of standardized means for performing the testing in liquid medium. We developed an assay using solid medium to identify interactions between antibiotics and phages for gram-positive and gram-negative bacteria. We modeled the interactions between antibiotics and phages on solid medium, and then tested multiple replicates of vancomycin-resistant Enterococcus (VRE) and Stenotrophomonas in the assay. For each organism, we identified synergy between different phage and antibiotic combinations. The development of this solid media assay for assessing synergy between phages and antibiotics will better inform the use of these combinations in the treatment of ARB infections.

A simple solid media assay for detection of synergy between bacteriophages and antibiotics.

The emergence of antibiotic resistant bacteria (ARB) has necessitated the development of alternative therapies to deal with this global threat. Bacteriophages (viruses that target bacteria) that kill ARB are one such alternative. While phages have been used clinically for decades with inconsistent results, a number of recent advances in phage selection, propagation and purification have enabled a reevaluation of their utility in contemporary clinical medicine. In most phage therapy cases, phages are administered in combination with antibiotics to ensure that patients receive the standard-of-care treatment. Some phages may work cooperatively with antibiotics to eradicate ARB, as often determined using non-standardized broth assays. We sought to develop a solid media-based assay to assess cooperativity between antibiotics and phages to offer a standardized platform for such testing. We modeled the interactions that occur between antibiotics and phages on solid medium to measure additive, antagonistic, and synergistic interactions. We then tested the method using different bacterial isolates, and identified a number of isolates where synergistic interactions were identified. These interactions were not dependent on the specific organism, phage family, or antibiotic used. A priori susceptibility to the antibiotic or the specific phage were not requirements to observe synergistic interactions. Our data also confirm the potential for the restoration of vancomycin to treat Vancomycin Resistant Enterococcus (VRE) when used in combination with phages. Solid media assays for the detection of cooperative interactions between antibiotics and phages can be an accessible technique adopted by clinical laboratories to evaluate antibiotic and phage choices in phage therapy.

Epidemiologic, clinical, and serum markers may improve discrimination between bacterial and viral etiologies of childhood pneumonia.

Background: Discrimination of bacterial and viral etiologies of childhood community-acquired pneumonia (CAP) is often challenging. Unnecessary antibiotic administration exposes patients to undue risks and may engender antimicrobial resistance. This study aimed to develop a prediction model using epidemiological, clinical and laboratory data to differentiate between bacterial and viral CAP. Methods: Data from 155 children with confirmed bacterial or mixed bacterial and viral infection (N = 124) and viral infection (N = 31) were derived from a comprehensive assessment of causative pathogens [Partnerships for Enhanced Engagement in Research-Pneumonia in Pediatrics (PEER-PePPeS)] conducted in Indonesia. Epidemiologic, clinical and biomarker profiles (hematology and inflammatory markers) were compared between groups. The area under the receiver operating characteristic curve (AUROC) for varying biomarker levels was used to characterize performance and determine cut-off values for discrimination of bacterial and mixed CAP versus viral CAP. Diagnostic predictors of bacterial and mixed CAP were assessed by multivariate logistic regression. Results: Diarrhea was more frequently reported in bacterial and mixed CAP, while viral infections more frequently occurred during Indonesia's rainy season. White blood cell counts (WBC), absolute neutrophil counts (ANC), neutrophil-lymphocyte ratio (NLR), C-reactive protein (CRP), and procalcitonin (PCT) were significantly higher in bacterial and mixed cases. After adjusting for covariates, the following were the most important predictors of bacterial or mixed CAP: rainy season (aOR 0.26; 95% CI 0.08-0.90; p = 0.033), CRP ≥5.70 mg/L (aOR 4.71; 95% CI 1.18-18.74; p = 0.028), and presence of fever (aOR 5.26; 95% CI 1.07-25.91; p = 0.041). The model assessed had a low R-squared (Nagelkerke R2 = 0.490) but good calibration (p = 0.610 for Hosmer Lemeshow test). The combination of CRP and fever had moderate predictive value with sensitivity and specificity of 62.28 and 65.52%, respectively. Conclusion: Combining clinical and laboratory profiles is potentially valuable for discriminating bacterial and mixed from viral pediatric CAP and may guide antibiotic use. Further studies with a larger sample size should be performed to validate this model.

Human Dectin-1 deficiency impairs macrophage-mediated defense against phaeohyphomycosis.

Subcutaneous phaeohyphomycosis typically affects immunocompetent individuals following traumatic inoculation. Severe or disseminated infection can occur in CARD9 deficiency or after transplantation, but the mechanisms protecting against phaeohyphomycosis remain unclear. We evaluated a patient with progressive, refractory Corynespora cassiicola phaeohyphomycosis and found that he carried biallelic deleterious mutations in CLEC7A encoding the CARD9-coupled, ß-glucan-binding receptor, Dectin-1. The patient's PBMCs failed to produce TNF-α and IL-1ß in response to ß-glucan and/or C. cassiicola. To confirm the cellular and molecular requirements for immunity against C. cassiicola, we developed a mouse model of this infection. Mouse macrophages required Dectin-1 and CARD9 for IL-1ß and TNF-α production, which enhanced fungal killing in an interdependent manner. Deficiency of either Dectin-1 or CARD9 was associated with more severe fungal disease, recapitulating the human observation. Because these data implicated impaired Dectin-1 responses in susceptibility to phaeohyphomycosis, we evaluated 17 additional unrelated patients with severe forms of the infection. We found that 12 out of 17 carried deleterious CLEC7A mutations associated with an altered Dectin-1 extracellular C-terminal domain and impaired Dectin-1-dependent cytokine production. Thus, we show that Dectin-1 and CARD9 promote protective TNF-α- and IL-1ß-mediated macrophage defense against C. cassiicola. More broadly, we demonstrate that human Dectin-1 deficiency may contribute to susceptibility to severe phaeohyphomycosis by certain dematiaceous fungi.

Epidemiology of community-acquired pneumonia among hospitalised children in Indonesia: a multicentre, prospective study.

OBJECTIVE: To identify aetiologies of childhood community-acquired pneumonia (CAP) based on a comprehensive diagnostic approach. DESIGN: 'Partnerships for Enhanced Engagement in Research-Pneumonia in Paediatrics (PEER-PePPeS)' study was an observational prospective cohort study conducted from July 2017 to September 2019. SETTING: Government referral teaching hospitals and satellite sites in three cities in Indonesia: Semarang, Yogyakarta and Tangerang. PARTICIPANTS: Hospitalised children aged 2-59 months who met the criteria for pneumonia were eligible. Children were excluded if they had been hospitalised for >24 hours; had malignancy or history of malignancy; a history of long-term (>2 months) steroid therapy, or conditions that might interfere with compliance with study procedures. MAIN OUTCOMES MEASURES: Causative bacterial, viral or mixed pathogen(s) for pneumonia were determined using microbiological, molecular and serological tests from routinely collected specimens (blood, sputum and nasopharyngeal swabs). We applied a previously published algorithm (PEER-PePPeS rules) to determine the causative pathogen(s). RESULTS: 188 subjects were enrolled. Based on our algorithm, 48 (25.5%) had a bacterial infection, 31 (16.5%) had a viral infection, 76 (40.4%) had mixed bacterial and viral infections, and 33 (17.6%) were unable to be classified. The five most common causative pathogens identified were Haemophilus influenzae non-type B (N=73, 38.8%), respiratory syncytial virus (RSV) (N=51, 27.1%), Klebsiella pneumoniae (N=43, 22.9%), Streptococcus pneumoniae (N=29, 15.4%) and Influenza virus (N=25, 13.3%). RSV and influenza virus diagnoses were highly associated with Indonesia's rainy season (November-March). The PCR assays on induced sputum (IS) specimens captured most of the pathogens identified in this study. CONCLUSIONS: Our study found that H. influenzae non-type B and RSV were the most frequently identified pathogens causing hospitalised CAP among Indonesian children aged 2-59 months old. Our study also highlights the importance of PCR for diagnosis and by extension, appropriate use of antimicrobials. TRAIL REGISTRATION NUMBER: NCT03366454.

Case Commentary: Long-Term Fosmanogepix Use in a Transplant Recipient with Disseminated Aspergillosis Caused by Azole-Resistant Aspergillus calidoustus.

Aspergillus calidoustus is an emerging, azole-resistant, cryptic Aspergillus species in immunosuppressed patients that often features extrapulmonary involvement and carries high mortality. The case presented by J. F. Camargo, R. Jabr, A. D. Anderson, L. Lekakis, et al. (Antimicrob Agents Chemother 66:e02206-21, 2022, https://doi.org/10.1128/aac.02206-21) describes a transplant recipient with disseminated A. calidoustus infection who was successfully treated with surgical source control, tapering of immunosuppression, and long-term, combination antifungal treatment that included the first-in-class fosmanogepix, which targets fungal mannoprotein trafficking and anchoring.

Approach to Identifying Causative Pathogens of Community-Acquired Pneumonia in Children Using Culture, Molecular, and Serology Tests.

Determining the causative pathogen(s) of community-acquired pneumonia (CAP) in children remains a challenge despite advances in diagnostic methods. Currently available guidelines generally recommend empiric antimicrobial therapy when the specific etiology is unknown. However, shifts in epidemiology, emergence of new pathogens, and increasing antimicrobial resistance underscore the importance of identifying causative pathogen(s). Although viral CAP among children is increasingly recognized, distinguishing viral from bacterial etiologies remains difficult. Obtaining high quality samples from infected lung tissue is typically the limiting factor. Additionally, interpretation of results from routinely collected specimens (blood, sputum, and nasopharyngeal swabs) is complicated by bacterial colonization and prolonged shedding of incidental respiratory viruses. Using current literature on assessment of CAP causes in children, we developed an approach for identifying the most likely causative pathogen(s) using blood and sputum culture, polymerase chain reaction (PCR), and paired serology. Our proposed rules do not rely on carriage prevalence data from controls. We herein share our perspective in order to help clinicians and researchers classify and manage childhood pneumonia.

Dacron swab and PBS are acceptable alternatives to flocked swab and viral transport media for SARS-CoV-2.

Nasopharyngeal flocked swabs placed in viral transport media (VTM) are the preferred collection methodology for respiratory virus testing. Due to the rapid depletion of available reagents and swabs, we have validated an alternative swab placed in phosphate-buffered saline (PBS) for use in respiratory virus testing in a SARS-CoV-2 real-time polymerase chain reaction assay and a multiplexed respiratory virus panel. We collected nasopharyngeal (NP) swabs and oropharyngeal (OP) swabs from 10 healthy volunteers. Flocked swabs were placed in VTM and alternative swabs in PBS. In this feasibility study, we show that NP collection is better for detection of human material than OP collection, as measured by significantly lower RNase P gene cycle threshold values, and that a Dacron polyester swab in PBS shows equivalent detection of SARS-CoV-2 and RSV to a flocked swab in VTM in contrived specimens. Diluted SARS-CoV-2-positive patient specimens are detectable for up to 72 h at 4 °C.

Mfge8 is critical for mammary gland remodeling during involution.

Apoptosis is a critical process in normal mammary gland development and the rapid clearance of apoptotic cells prevents tissue injury associated with the release of intracellular antigens from dying cells. Milk fat globule-EGF-factor 8 (Mfge8) is a milk glycoprotein that is abundantly expressed in the mammary gland epithelium and has been shown to facilitate the clearance of apoptotic lymphocytes by splenic macrophages. We report that mice with disruption of Mfge8 had normal mammary gland development until involution. However, abnormal mammary gland remodeling was observed postlactation in Mfge8 mutant mice. During early involution, Mfge8 mutant mice had increased numbers of apoptotic cells within the mammary gland associated with a delay in alveolar collapse and fat cell repopulation. As involution progressed, Mfge8 mutants developed inflammation as assessed by CD45 and CD11b staining of mammary gland tissue sections. With additional pregnancies, Mfge8 mutant mice developed progressive dilatation of the mammary gland ductal network. These data demonstrate that Mfge8 regulates the clearance of apoptotic epithelial cells during mammary gland involution and that the absence of Mfge8 leads to inflammation and abnormal mammary gland remodeling.