Eosinophilia in Infectious Diseases.

Paragonimus and paragonimiasis in Asia: An update.

Cough Due to TB and Other Chronic Infections: CHEST Guideline and Expert Panel Report

Paragonimiasis caused by lung flukes (genus Paragonimus) is a neglected disease occurring in Asia, Africa and the Americas. The genus is species-rich, ancient and widespread. Genetic diversity is likely to be considerable, but investigation of this remains confined to a few populations of a few species. In recent years, studies of genetic diversity have moved from isoenzyme analysis to molecular phylogenetic analysis based on selected DNA sequences. The former offered better resolution of questions relating to allelic diversity and gene flow, whereas the latter is more suitable for questions relating to molecular taxonomy and phylogeny. A picture is emerging of a highly diverse taxon of parasites, with the greatest diversity found in eastern and southern Asia where ongoing speciation might be indicated by the presence of several species complexes. Diversity of lung flukes in Africa and the Americas is very poorly sampled. Functional molecules that might be of value for immunodiagnosis, or as targets for medical intervention, are of great interest. Characterisation of these from Paragonimus species has been ongoing for a number of years. However, the imminent release of genomic and transcriptomic data for several species of Paragonimus will dramatically increase the rate of discovery of such molecules, and illuminate their diversity within and between species.

Gene diversity and genetic variation in lung flukes (genus Paragonimus)

Background: Paragonimiasis is a food-borne trematode infection acquired by eating raw or undercooked crustaceans. It is a major public health problem in the far East, but it also occurs in South Asia, Africa, and in the Americas. Paragonimus worms cause chronic lung disease with cough, fever and hemoptysis that can be confused with tuberculosis or other non-parasitic diseases. Treatment is straightforward, but diagnosis is often delayed due to a lack of reliable parasitological or serodiagnostic tests. Hence, the purpose of this study was to use a systems biology approach to identify key parasite proteins that may be useful for development of improved diagnostic tests. Methodology/Principal Findings: The transcriptome of adult Paragonimus kellicotti was sequenced with Illumina technology. Raw reads were pre-processed and assembled into 78,674 unique transcripts derived from 54,622 genetic loci, and 77,123 unique protein translations were predicted. A total of 2,555 predicted proteins (from 1,863 genetic loci) were verified by mass spectrometric analysis of total worm homogenate, including 63 proteins lacking homology to previously characterized sequences. Parasite proteins encoded by 321 transcripts (227 genetic loci) were reactive with antibodies from infected patients, as demonstrated by immunoaffinity purification and high-resolution liquid chromatography-mass spectrometry. Serodiagnostic candidates were prioritized based on several criteria, especially low conservation with proteins in other trematodes. Cysteine proteases, MFP6 proteins and myoglobins were abundant among the immunoreactive proteins, and these warrant further study as diagnostic candidates. Conclusions: The transcriptome, proteome and immunolome of adult P. kellicotti represent a major advance in the study of Paragonimus species. These data provide a powerful foundation for translational research to develop improved diagnostic tests. Similar integrated approaches may be useful for identifying novel targets for drugs and vaccines in the future.

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Systems biology studies of adult paragonimus lung flukes facilitate the identification of immunodominant parasite antigens.

Background:Pulmonary tuberculosis (PTB) and paragonimiasis remain as health problems in certain areas in the Philippines. Both share similar clinical manifestations, which include chronic productive cough, hemoptysis, dyspnea, fever, weight loss, and night sweats. This study aimed to determine the prevalence of PTB, paragonimiasis, and co-infections in Zamboanga del Norte, Philippines.Methods:This study was conducted in selected villages in two municipalities in Zamboanga del Norte. Patients with chronic cough were interviewed, examined, and requested to submit two sputum samples which were processed using Ziehl–Neelsen method to detect acid-fast bacilli (AFB), and NaOH concentration technique for the detection of Paragonimus ova.Results:A total of 836 patients submitted sputum samples for examination. Prevalence was 6·7% (2·5–12·7%) for paragonimiasis and 1·9% (0·9–6·3%) for PTB. Co-infection rate was 0·3%, with two identified cases. Positivity rates for males and females were 9·6 and 5·8% for pa...

Integrated surveillance of pulmonary tuberculosis and paragonimiasis in Zamboanga del Norte, the Philippines

A 48-year-old man presented with a non-healing wound on his left foot after stepping on a nail. He self-medicated with amoxicillin, but the wound progressed prompting consult. On examination, his left foot was diffusely swollen with surrounding erythema, areas of gangrene, foul-smelling purulent discharge and subcutaneous emphysema. He was managed as a case of necrotising fasciitis and underwent emergent amputation. Three days after amputation, he developed a sudden and progressive blurring of vision, swelling and conjunctival erythema, with purulent discharge and the presence of hypopyon on the left eye. He was then managed as a case of endophthalmitis of the left eye and underwent pars plana vitrectomy. All cultures (blood, tissue and vitreous fluid) grew pan-susceptible hypermucoviscous Klebsiella pneumoniae, with positive string tests and confirmed by multilocus gene sequencing and sequence type analysis. He gradually improved with intravenous antibiotics, but only regained light perception in the left eye.

Hypermucoviscous capsular 1 (K1) serotype Klebsiella pneumoniae necrotising fasciitis and metastatic endophthalmitis.

Improving paragonimiasis surveillance, which is crucial for disease control, requires adopting new tools and techniques useful in mapping endemic areas. This study aimed to (1) develop a questionna...

Identification of suspected paragonimiasis-endemic foci using a questionnaire and detection of Paragonimus ova using the Ziehl-Neelsen technique in Zamboanga Region, the Philippines.

Body: Noroviruses are one of the leading causes of acute gastroenteritis (AGE), characterized by diarrheal illness and/or vomiting and accounting for approximately one-fifth of AGE cases worldwide. Despite its ubiquity, little is known about the effects of norovirus-associated AGE on development of gut microbiomes in infants, which is hypothesized to play a role in the development of robust immune responses. Using a shotgun metagenomics-based approach to detect and quantify changes in the taxonomic and functional diversity incurred by AGE-induced disturbance, we sequenced stools at time points before, during, and after the first norovirus-associated AGE occurring in five children enrolled in a birth cohort study conducted in León, Nicaragua. Amongst the children, the taxonomic and functional diversity of microbiomes increased substantially during the first symptomatic norovirus episode, with a dominance of Gammaproteobacteria. After recovery from norovirus AGE, there was a return of many of the probiotic microbial taxa typical of breast-feeding infants profiled prior to the norovirus AGE episode. Most notably, our findings demonstrate a temporary disturbance to the microbiome during norovirus-associated AGE that did not alter the development of healthy microbiomes amongst the immunologically naïve children in this birth cohort. The metagenomic approach presented here should be broadly applicable to other AGE-associated pathogens to provide foundational data important for developing probiotic treatments and vaccines. Abstract Body: Objectives: Candida auris is a multidrug-resistant pathogenic fungus, capable of causing invasive infections with high mortality. Despite intense efforts to understand how this pathogen rapidly emerged and spread to healthcare facilities worldwide, the environmental reservoir of C. auris remains unknown. Environmental surveillance aimed at identifying origins for emerging pathogens requires extensive sampling that is often not feasible. However, as metagenomic studies expand to encompass more environments, it is now possible to flip the traditional paradigm of collecting and screening individual samples for specific pathogen sequences and instead leverage big data repositories to identify all Abstract Body: Members of the bacterial genus Streptomyces (phylum Actinobacteria) are best known as major bacterial producers of antibiotics and other useful compounds commonly used in human medicine, animal health, and agriculture. They are ubiquitous in nature, often inhabiting diverse soil environments. They are also often associated with animals, yet their diversity in these habitats and the evolutionary processes that shape such diversity are often poorly understood. Here, we investigated the contributions of homologous recombination and selection on the core genome of 73 previously published Streptomyces genomes from bats. Of the 1141 core genes, calculation of the pairwise homoplasy index revealed 924 genes that did not show evidence of recombination and 217 genes that have experienced recombination. We also estimated the ratio of non-synonymous to synonymous polymorphisms (dN/dS) of each core gene and found 116 that show evidence of positive selection. Combining the two analyses, we found six frequently recombining genes that also exhibit positive selection: ettA3 , gcvT1 , guaA , lpdA , nrdZ , sdaA . We also identified 207 frequently recombining genes that also exhibit negative (purifying) selection. We discuss the functions of these genes in the context of host-microbe interactions and natural drug discovery efforts. Overall, these results indicate that genes acquired through recombination can be maintained in recipient organisms through selection, thereby facilitating the adaptation to their bat hosts. Abstract Body: We aimed to determine the relationships among milk microbiota, milk components (lactose, oligosaccharides (MO) and fatty acids (MFA)), and somatic cell counts (SCC), in lactating Holstein cows. Raw milk samples were collected from cows at three timepoints, ranging from early to late lactation. Milk microbial community was determined by 16S rRNA amplicon sequencing, lactose abundance by Fourier transform infrared spectroscopy, MO by nano-liquid chromatography quadrupole time-of-flight tandem mass spectrometry, MFA by fatty acid methyl esters analysis by gas chromatography with flame ionization detection, and SCC (as a measure of mammary inflammation) by flow cytometry. Abstract Body: Human noroviruses (HuNoV) are a leading cause of acute gastroenteritis across all ages in the United States. Genetic differences in histo-blood group antigen (HBGA) expression drives susceptibility to different HuNoV strains. While antibodies that block HuNoV binding to HBGAs correlate with protection from illness, whether other host and microbial factors influence HuNoV gastroenteritis is unknown. We used samples from a human experimental infection study to determine whether whole genome shotgun (WGS) metagenomic profiles, including taxonomy and enriched pathways, can predict the risk of developing gastroenteritis in genetically susceptible individuals. We profiled 604 stool samples from 55 healthy adults who were challenged with the prototype HuNoV Norwalk virus (GI.1) or placebo using WGS metagenomics. Samples collected pre-challenge and at multiple visits (day 2-56) post-challenge were tested. Of 41 genetically susceptible persons, 20 were infected and 13 developed symptoms of acute gastroenteritis (diarrhea and/or vomiting). Machine learning classification was performed pre- and post-challenge on a combined set of clades and pathways via RandomForest (RF) and Support Vector Machines (SVM) to build models predicting acute gastroenteritis. We used cross-validation, dividing our samples 80% for training and 20% for testing, using the Area Under the Curve (AUC) metric. Fifty iterations were performed for each model and median AUC was reported. Each sample was considered independently. WGS analysis identified 603 clades and 533 pathways. In pre-challenge profiles of individuals who received high dose GI.1 inoculum (4800 or 48 genome equivalents, GE), RF models achieved AUC=0.99; Dorea formicigenerans and Gemmiger formicilis were more abundant in persons with acute gastroenteritis. In those who received low doses of GI.1 inoculum (4.8 or 0.48 GE), SVM achieved AUC=0.99, with higher abundance of Streptococcus vestibularis and Denitrobacterium detoxificans in persons with acute gastroenteritis. In post-infection profiles, RF and SVM achieved AUC=0.99 for both high and low dose groups, revealing higher abundance of Eubacterium hallii , D. formicigenerans and G. formicilis in the high dose group and higher abundance of S. vestibularis and Dialister invisus in the low dose group. Despite a relatively low sample size and considering each sample independently, we built effective models to predict acute gastroenteritis with GI.1 HuNoV. Identifying key microbial species associated with gastroenteritis will broaden our understanding of HuNoV pathophysiology and aid the design of potential therapies. Chronic obstructive pulmonary disease (COPD) is a common condition associated with significant morbidity and mortality as well as increased health care costs. Smoke exposure has been shown to be a causative factor in its development. However, some patients with a heavy smoking history may not develop physiological evidence of COPD. In this study we sought to investigate whether lower airway dysbiosis occurs in early-stage COPD and contributes to the pathogenesis of the disease. To explore this, we recruited 57 patients with a greater than 20 pack year smoking history to perform an in-depth microbiome analysis. Of the 57 patients, 26 had physiological evidence of COPD and the remaining were grouped as smoker controls. All patients had a bronchoscopy performed where lower airway (BAL), upper airway (UA) and bronchoscopy control (BKG) samples were collected. Samples were analyzed by 16S rRNA gene sequencing, whole genome sequencing, RNA metatranscriptome and host RNA transcriptome sequencing. In addition, a mouse model was used to evaluate the different contributions that smoke and dysbiosis have on lower airway inflammatory injury. Abstract The ability to field genome-sensing technologies in distributed, austere locations changes the nature of the public health practitioner’s relationship with extant biological information present in the environment. Determining the presence of active pathogens (naturally-occurring or synthetic/weaponized) is a key advantage for biosurveillance and biodefense applications. Accessing direct sequence-level information, as opposed to indirect detection of sequence-level signatures via amplification technologies (e.g., PCR), confers distinct bio-intelligence benefits. These benefits are most acutely realized when the active biological threat is an unknown, emergent entity without any comparable reference in genome repositories. Synthetically-engineered chimeric biothreats are poised to become a more pressing biodefense concern, underscoring the limitations to pre-designed assay technology and the need for direct sequence-level data sensing. Currently, nanopore-based sequencing platforms are the most developed technologies that could meet this need in a tactical or field-forward environment, outside of traditional laboratories and more proximal to the point-of-sampling. However, the maturity of these platforms for the rigors of tactical scenarios are still evolving to the required levels. Here, we field-test Oxford Nanopore's MinION sequencing device, embedded in a ruggedized tactical laboratory platform (Mercury Lab, Figure 1), for the purposes of un-biased sequence-level surveillance and detection of pathogen signatures directly on site, from arthropod samples found in the testing environment. The field site -- St. Catherines Island -- is a barrier island off the coast of the U.

Manuel Jorge

Papers

Integrated surveillance of pulmonary tuberculosis and paragonimiasis in Zamboanga del Norte, the Philippines

Hypermucoviscous capsular 1 (K1) serotype Klebsiella pneumoniae necrotising fasciitis and metastatic endophthalmitis.

Identification of suspected paragonimiasis-endemic foci using a questionnaire and detection of Paragonimus ova using the Ziehl-Neelsen technique in Zamboanga Region, the Philippines.

Leveraging Metagenomic Data from SRA for finding Candida auris reservoirs