Biology subjects

Qadri, F.

Publications and source records attributed to Qadri, F..

27 records · Page 2Linked to original sources

bioRxiv2020

Contribution of noncanonical antigens to virulence and adaptive immunity in human infection with enterotoxigenic E. coli

Enterotoxigenic E. coli (ETEC) contribute significantly to the substantial burden of infectious diarrhea among children living in low and middle income countries. In the absence of a vaccine for ETEC, children succumb to acute dehydration as well as non-diarrheal sequelae related to these infections including malnutrition. The considerable diversity of ETEC genomes has complicated canonical vaccine development approaches focused on a subset of antigens known as colonization factors (CFs). To identify additional conserved immunogens, we mined genomic sequences of 89 ETEC isolates, bioinformatically selected potential surface-exposed pathovar-specific antigens conserved in more than 40% of the genomes (n=118), and assembled the representative proteins onto microarrays, complemented with known or putative colonization factor subunit molecules (n=52), and toxin subunits to interrogate samples from individuals with acute symptomatic ETEC infections. Surprisingly, in this open-aperture approach, we found that immune responses were largely constrained to a small number of antigens including individual colonization factor antigens and EtpA, an extracellular adhesin. In a Bangladeshi cohort of naturally infected children < 2 years of age, both EtpA and a second noncanonical antigen, EatA, elicited significant serologic responses that were associated with protection from symptomatic illness. In addition, children infected with ETEC isolates bearing either etpA or eatA genes were significantly more likely to develop symptomatic disease. These studies support a role for more recently discovered noncanonical antigens in virulence and the development of adaptive immune responses during ETEC infections, findings that may inform vaccine design efforts to complement existing approaches.

microbiology↗

bioRxiv2020

A combination of metagenomic and cultivation approaches reveals hypermutator phenotypes within Vibrio cholerae infected patients

Vibrio cholerae can cause a range of symptoms, ranging from severe diarrhea to asymptomatic infection. Previous studies using whole genome sequencing (WGS) of multiple bacterial isolates per patient showed that V. cholerae can evolve modest genetic diversity during symptomatic infection. To further explore the extent of V. cholerae within-host diversity, we applied culture-based WGS and metagenomics to a cohort of both symptomatic and asymptomatic cholera patients from Bangladesh. While metagenomics allowed us to detect more mutations in symptomatic patients, WGS of cultured isolates was necessary to detect V. cholerae diversity in asymptomatic carriers, likely due to their low V. cholerae load. Using both metagenomics and isolate WGS, we report three lines of evidence that V. cholerae hypermutators evolve within patients. First, we identified nonsynonymous in V. cholerae DNA repair genes in five out of 11 patient metagenomes sequenced with sufficient coverage of the V. cholerae genome, and in one of three patients with isolate genomes sequenced. Second, mutations in DNA repair genes tended to be accompanied by an excess of intrahost single nucleotide variants (iSNVs). Third, these iSNVs were enriched in transversion mutations, a known hallmark of hypermutator phenotypes. While hypermutators appeared to generate mostly selectively neutral mutations, non-mutators showed signs of convergent mutation across multiple patients, suggesting V. cholerae adaptation within hosts. Our results highlight the power and limitations of metagenomics combined with isolate sequencing to characterize within-patient diversity in acute V. cholerae infections, while providing evidence for hypermutator phenotypes within cholera patients. IMPORTANCEPathogen evolution within patients can impact phenotypes such as drug resistance and virulence, potentially affecting clinical outcomes. V. cholerae infection can result in life-threatening diarrheal disease, or asymptomatic infection. Here we describe whole-genome sequencing of V. cholerae isolates and culture-free metagenomic sequencing from stool of symptomatic cholera patients and asymptomatic carriers. Despite the typically short duration of cholera, we found evidence for adaptive mutations in the V. cholerae genome that occur independently and repeatedly within multiple symptomatic patients. We also identified V. cholerae hypermutator phenotypes within several patients, which appear to generate mainly neutral or deleterious mutations. Our work sets the stage for future studies of the role of hypermutators and within-patient evolution in explaining the variation from asymptomatic carriage to symptomatic cholera.

microbiology↗

bioRxiv2020

Humans surviving cholera develop antibodies against Vibrio cholerae O-specific polysaccharide that inhibit pathogen motility

The mechanism of protection against cholera afforded by previous illness or vaccination is currently unknown. We have recently shown that antibodies targeting O-specific polysaccharide (OSP) of Vibrio cholerae correlate highly with protection against cholera. V. cholerae is highly motile and possesses a flagellum sheathed in O-specific polysaccharide (OSP), and motility of V. cholerae correlates with virulence. Using high speed video microscopy, and building upon previous animal-related work, we demonstrate that sera, polyclonal antibody fractions, and OSP-specific monoclonal antibodies recovered from humans surviving cholera block V. cholerae motility at both subagglutinating and agglutinating concentrations. This anti-motility effect is reversed by pre-adsorbing sera and polyclonal antibody fractions with purified OSP; and is associated with OSP-specific but not flagellin-specific monoclonal antibodies. F[ab] fragments of OSP-specific polyclonal antibodies do not inhibit motility, suggesting a requirement for antibody-mediated crosslinking in motility inhibition. We show that OSP-specific antibodies do not directly affect V. cholerae viability, but that OSP-specific monoclonal antibody highly protects against death in the murine cholera model. We used in vivo competitive index studies to demonstrate that OSP-specific antibodies impede colonization and survival of V. cholerae in intestinal tissues, and that this impact is motility-dependent. Our findings suggest that the impedance of motility by antibodies targeting V. cholerae OSP contributes to protection against cholera. IMPORTANCECholera is a severe dehydrating illness of humans caused by Vibrio cholerae. V. cholerae is a highly motile bacterium that has a single flagellum covered in lipopolysaccharide (LPS) displaying O-specific polysaccharide (OSP), and V. cholerae motility correlates with its ability to cause disease. The mechanisms of protection against cholera are not well understood; however, since V. cholerae is a non-invasive intestinal pathogen, it is likely that antibodies that bind the pathogen or its products in the intestinal lumen contribute to protection from infection. Here, we demonstrate that OSP-specific antibodies isolated from humans surviving cholera in Bangladesh inhibit V. cholerae motility and are associated with protection against challenge in a motility-dependent manner.

microbiology↗

bioRxiv2020

Association of poor virus specific immunoglobulin G antibodyresponses with higher viral load is seen in Bangladeshi pregnantwomen having acute Hepatitis E Genotype 1 infection

Although Hepatitis E viral illness is usually self-limiting, higher rates of morbidity and mortality are frequently observed during pregnancy in South Asian countries including Bangladesh. Of the four common variants, hepatitis E virus genotype 1 is mainly prevalent in South Asian countries. Pregnant women usually suffer from a state of immunosuppression. It is yet to be known whether virus specific immunoglobulin G (IgG) immune responses have any association with the vulnerability of pregnant women to acute hepatitis with E virus. The study aimed to compare the viral load and IgG responses of hepatitis E-infected pregnant women with that of non-pregnant women with same infection. Real Time -quantitative reverse transcription Polymerase Chain Reaction and Sanger sequencing were performed to determine the viral load and genotype, respectively, whereas Enzyme Linked Immunosorbent Assay method was used to determine hepatitis E virus specific serum IgG antibody index along with IgG avidity index. Although significant negative correlations were observed between log viral copy number and log IgG antibody index in the late acute phases of jaundice for both pregnant (r= -0.7971, p=0.0002) and non-pregnant women (r= -0.9117, p=0.0002), serum log viral copy number of pregnant women was significantly higher than that of the non-pregnant counterpart (p=0.0196) in the late acute stage of jaundice. In addition, log hepatitis E virus IgG antibody index of pregnant women was significantly lower than the non-pregnant women in the late phase of jaundice induced by hepatitis E virus (p=0.0303). Moreover, pregnant women with acute hepatitis E had higher cross-reactive IgG than in the non-pregnant women (p=0.0017). All the patients got infected with hepatitis E virus were in Genotype 1 variety. The study demonstrates that virus-specific poor IgG responses might be responsible for vulnerability of pregnant women to acute hepatitis with hepatitis E virus. Author SummaryAcute hepatitis caused by hepatitis E virus (HEV) Genotype 1 is a public health problem in Asian countries and especially it poses a potential health threat to pregnant women causing 19% to 25% mortality, particularly in South Asian countries including Bangladesh. The study aimed to explore whether HEV IgG immune responses were compromised during pregnancy, which might contribute to higher viral load and disease severity. Accordingly, pregnant and non-pregnant women with acute hepatitis (clinically presented with nausea, loss of appetite and /or jaundice) were enrolled from different tertiary care hospitals in Dhaka city. All these patients were screened and hepatitis E were differentiated from other hepatitis (caused by A, B, C) using Enzyme Linked Immunosorbent Assay (ELISA) methods. HEV IgG antibody/avidity indices and viral loads were measured using ELISA and real time quantitative polymerase chain reaction (RT-qPCR), respectively. The study showed that pregnant women with acute hepatitis E had lower IgG indices with higher viral load than their non-pregnant counterpart. Overall, the study revealed that virus-specific poor IgG responses might render pregnant women vulnerable to acute hepatitis E of varying degree of severity which might be associated with higher viral load.

immunology↗

bioRxiv2020

Situation of antibiotic resistance in Bangladesh and its association with resistance genes for horizontal transfer

The study investigated the spectrum of antibiotic resistance and the associated genes for aminoglycoside, macrolide and ESBL class of antibiotics using clinical isolates. A total of 430 preserved bacterial strains (Acinetobacter baumannii, n= 20; Pseudomonas aeruginosa, n= 26; Klebsiella pneumoniae, n= 42; E.coli, n= 85; Staphylococcus aureus, n= 84; Salmonella Typhi, n= 82; Enterococcus spp., n= 27; Streptococcus pneumoniae, n= 36 and CNS, n= 28) were examined. The strains were isolated from patients admitted to various tertiary hospitals of Dhaka city between 2015 and 2019 with either acute respiratory infections, wound infections, typhoid fever or diarrhea. The isolates were reconfirmed by appropriate microbiological and biochemical methods. Antimicrobial susceptibility tests were done using Kirby-Bauer disk diffusion approach. PCR amplification using resistance gene-specific primers for aminoglycoside, macrolide and ESBL class of antibiotics was done and the amplified products were confirmed by Sanger sequencing. Of the total isolates, 53% came out as MDR with 96.6% of E. coli and 90% of Staphylococcus aureus. There was a year-wise gradual increase of MDR isolates from 2015-2018 and by 2019 the increase in MDR isolates became almost 2-fold compared to 2015. Among the five ESBL genes investigated, CTXM-1 came out as the most prevalent (63%) followed by NDM-1 (22%) and E. coli isolates were the predominant reservoir of these genes. ErmB (55%) was the most frequently detected macrolide resistance gene, whereas aac(6)-Ib (35.44%) was the most prevalent aminoglycoside resistance gene and these genes were most prevalent in E. coli and P. aeruginosa isolates, respectively. CTXM-1 and ErmB (16.66%) were the most frequent partners of coexistence followed by CTXM-1 and aac(3)-II.

microbiology↗

bioRxiv2020

Predicting Vibrio cholerae infection and disease severity using metagenomics in a prospective cohort study

BackgroundSusceptibility to Vibrio cholerae infection is impacted by blood group, age, and pre-existing immunity, but these factors only partially explain who becomes infected. A recent study used 16S rRNA amplicon sequencing to quantify the composition of the gut microbiome and identify predictive biomarkers of infection with limited taxonomic resolution. MethodsTo achieve increased resolution of gut microbial factors associated with V. cholerae susceptibility and identify predictors of symptomatic disease, we applied deep shotgun metagenomic sequencing to a cohort of household contacts of patients with cholera. ResultsUsing machine learning, we resolved species, strains, gene families, and cellular pathways in the microbiome at the time of exposure to V. cholerae to identify markers that predict infection and symptoms. Use of metagenomic features improved the precision and accuracy of prediction relative to 16S sequencing. We also predicted disease severity, although with greater uncertainty than our infection prediction. Species within the genera Prevotella and Bifidobacterium predicted protection from infection, and genes involved in iron metabolism also correlated with protection. ConclusionOur results highlight the power of metagenomics to predict disease outcomes and suggest specific species and genes for experimental testing to investigate mechanisms of microbiome-related protection from cholera. SUMMARYCholera infection and disease severity can be predicted using metagenomic sequencing of the gut microbiome pre-infection in a prospective cohort, and suggests potentially protective bacterial species and genes.

microbiology↗

bioRxiv2019

Population structure and antimicrobial resistance patterns of Salmonella Typhi isolates in Bangladesh from 2004 to 2016

BackgroundMulti-drug resistant typhoid fever remains an enormous public health threat in low and middle-income countries. However, we still lack a detailed understanding of the epidemiology and genomics of S. Typhi in many regions. Here we have undertaken a detailed genomic analysis of typhoid in Bangladesh to unravel the population structure and antimicrobial resistance patterns in S. Typhi isolated in between 2004-2016.\n\nPrincipal findingsWhole genome sequencing of 202 S. Typhi isolates obtained from three study locations in urban Dhaka revealed a diverse range of S. Typhi genotypes and AMR profiles. The bacterial population within Dhaka were relatively homogenous with little stratification between different healthcare facilities or age groups. We also observed evidence of transmission of Bangladeshi genotypes with neighboring South Asian countries (India, Pakistan and Nepal) suggesting these are circulating throughout the region. This analysis revealed a decline in H58 (genotype 4.3.1) isolates from 2011 onwards, coinciding with a rise in a diverse range of non-H58 genotypes and a simultaneous rise in isolates with reduced susceptibility to fluoroquinolones, potentially reflecting a change in treatment practices. We identified a novel S. Typhi genotype, subclade 3.3.2 (previously defined only to clade level, 3.3), which formed two localised clusters (3.3.2.Bd1 and 3.3.2.Bd2) associated with different mutations in the Quinolone Resistance Determining Region (QRDR) of gene gyrA.\n\nSignificanceOur analysis of S. Typhi isolates from Bangladesh isolated over a twelve year period identified a diverse range of AMR profiles and genotypes. The observed increase in non-H58 genotypes associated with reduced fluoroquinolone susceptibility may reflect a change in treatment practice in this region and highlights the importance of continued molecular surveillance to monitor the ongoing evolution of AMR in Bangladesh. We have defined new genotypes and lineages of Bangladeshi S. Typhi which will facilitate identification of these emerging AMR clones in future surveillance efforts.\n\nAuthor SummaryTyphoid fever, caused by Salmonella enterica serovar Typhi, is an acute and often life-threatening febrile illness in developing countries. Until recently, there have been limited studies focusing on the epidemiology and disease burden of typhoid in poor resource settings including Bangladesh. This study highlights the urgent need for sustained genomics based surveillance studies to monitor the population structure and ongoing evolution of AMR. Our data revealed a diverse range of S. Typhi genotypes and AMR patterns among 202 isolates collected from three urban areas in Dhaka. Moreover, we defined a new genotype, subclade 3.3.2 (previously typed only to clade level, 3.3) with two Bangladesh-localiased clades 3.3.2.Bd1 and 3.3.2.Bd2 showing reduced susceptibility to fluoroquinolones. Our data shows a significant increase in non-H58 genotypes carrying QRDR mutations from 2012 onwards, replacing MDR H58 genotypes. Our data suggest that a shift in treatment practice towards third generation cephalosporins to control typhoid might be beneficial, in addition to the introduction of vaccination programs and improvements in water sanitation and hygiene (WASH) in Bangladesh.

genomics↗

bioRxiv2018

Cognate T and B cell interaction and association of Follicular helper T cells with B cell responses in Vibrio cholerae O1 infected Bangladeshi adults

Vibrio cholerae O1 can cause life threatening diarrheal disease if left untreated. A long lasting immune response, producing 3-5 years of protection from subsequent, symptomatic disease following natural infection, is mediated by B cell mediated humoral immunity. T cells can play critical roles in inducing such immunity. However, the mechanism of T cell dependent B cell maturation and whether a key sub-population of T cells are involved is not well established in cholera. We hypothesized that a specific population of T cells, follicular helper T (Tfh) cells, are involved in B cell maturation following cholera; we used flow cytometry, culture and colorimetric assays to address this question. We found that V. cholerae infection induces significant increase in circulating Tfh cells expressing B cell maturation associated protein CD40L early in disease. The increased Tfh cells expressing CD40L recognize cholera toxin most prominently, with lessened responses to two antigens tested, V. cholerae membrane preparation (MP) and Vibrio cholerae cytolysin (VCC). We further showed that early induction of Tfh cells and CD40L was associated with later memory B cell responses to same antigens. Lastly, we demonstrated in vitro that Tfh cells isolated after cholera can stimulate class switching of cocultured, isolated B cells from patients with cholera, leading to production of the more durable IgG antibody isotype. These studies were conducted on circulating Tfh cells; future studies will be directed at examining role of Tfh cells during cholera directly in the gut mucosa of biopsied samples, at the single cell level if feasible.

immunology↗

bioRxiv2017

Vibrio cholerae genomic diversity within and between patients

Cholera is a severe, waterborne diarrheal disease caused by toxin-producing strains of the bacterium Vibrio cholerae. Comparative genomics has revealed \"waves\" of cholera transmission and evolution, in which clones are successively replaced over decades and centuries. However, the extent of V. cholerae genetic diversity within an epidemic or even within an individual patient is poorly understood. Here, we characterized V. cholerae genomic diversity at a micro-epidemiological level within and between individual patients from Bangladesh and Haiti. To capture within-patient diversity, we isolated multiple (8 to 20) V. cholerae colonies from each of eight patients, sequenced their genomes and identified point mutations and gene gain/loss events. We found limited but detectable diversity at the level of point mutations within hosts (zero to three single nucleotide variants within each patient), and comparatively higher gene content variation within hosts (at least one gain/loss event per patient, and up to 103 events in one patient). Much of the gene content variation appeared to be due to gain and loss of phage and plasmids within the V. cholerae population, with occasional exchanges between V. cholerae and other members of the gut microbiota. We also show that certain intra-host variants have phenotypic consequences. For example, the acquisition of a Bacteroides plasmid and nonsynonymous mutations in a sensor histidine kinase gene both reduced biofilm formation, an important trait for environmental survival. Together, our results show that V. cholerae is measurably evolving within patients, with possible implications for disease outcomes and transmission dynamics.\n\nAuthor SummaryVibrio cholerae is the etiological agent of cholera, a severe diarrheal disease endemic to Bangladesh and responsible for global outbreaks, including one ongoing in Haiti. Certain bacterial pathogens can evolve and diversify within the human host, often altering virulence and antibiotic resistance. However, most examples of within-host evolution have come from chronic infections, in which the pathogen has sufficient time to mutate and diversify, and little attention has been paid to more acute infections such as the one caused by V. cholerae. The goal of this study was to measure the extent of within-host evolution of V. cholerae within individual infected patients. By sequencing multiple bacterial isolates_from each of eight patients from Bangladesh and Haiti, we found that cholera patients can harbor a diverse population of V. cholerae. As expected for an acute infection, this diversity is limited, ranging from zero to three point mutations (single nucleotide variants) per patient. However, gene gain/loss events are more prevalent than point mutations, occurring in every single patient, and sometimes involving the transfer of dozens of genes on plasmids. Even if rare, point mutations and gene gain/loss events may be maintained by natural selection, and can alter clinically-and environmentally-relevant phenotypes such as biofilm formation. Therefore, within-patient evolution has the potential to impact clinical and epidemiological outcomes. Together, our results demonstrate that within-patient evolution may be a general feature of both acute and chronic infections, and that gene gain/loss may be an important but under-appreciated feature of within-host evolution.

microbiology↗