Biology subjects

Rahman, A.

Publications and source records attributed to Rahman, A..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

MLC1 alteration in iPSCs give rise to disease-like cellular vacuolation phenotype in the astrocyte lineage

BackgroundMegalencephalic leukoencephalopathy with subcortical cysts (MLC), a rare and progressive neurodegenerative disorder involving the white matter, is not adequately recapitulated by current disease models. Somatic cell reprogramming, along with advancements in genome engineering, may allow the establishment of in-vitro human models of MLC for disease modeling and drug screening. In this study, we utilized cellular reprogramming and gene-editing techniques to develop induced pluripotent stem cell (iPSC) models of MLC to recapitulate the cellular context of the classical MLC-impacted nervous system. MethodsSomatic cell reprogramming of peripheral patient-derived blood mononuclear cells (PBMCs) was used to develop iPSC models of MLC. CRISPR-Cas9 system-based genome engineering was also utilized to create the MLC1 knockout model of the disease. Directed differentiation of iPSCs to neural stem cells (NSCs) and astrocytes was performed in a 2D cell culture format, followed by various cellular and molecular biology approaches, to characterize the disease model. ResultsMLC iPSCs established by somatic cell reprogramming and genome engineering were well characterized for pluripotency. iPSCs were subsequently differentiated to disease-relevant cell types: neural stem cells (NSCs) and astrocytes. RNA sequencing profiling of MLC NSCs revealed a set of differentially expressed genes related to neurological disorders and epilepsy, a common clinical finding within MLC disease. This gene set can serve as a target for drug screening for the development of a potential therapeutic for this disease. Upon differentiation to the more disease relevant cell type-astrocytes, MLC-characteristic vacuoles were clearly observed, which were distinctly absent from controls. This emergence recapitulated a distinguishing phenotypic marker of the disease. ConclusionThrough the creation and analyses of iPSC models of MLC, our work addresses a critical need for relevant cellular models of MLC for use in both disease modeling and drug screening assays. Further investigation can utilize MLC iPSC models, as well as generated transcriptomic data sets and analyses, to identify potential therapeutic interventions for this debilitating disease.

cell biology↗

bioRxiv2024

GAGER: gene regulatory network assisted gene expression restoration

Gene regulatory networks are crucial for cellular function, and disruptions in transcription factor (TF) regulation often lead to diseases. However, identifying TFs to transition a source cell state to a desired target state remains challenging. We present a method to identify key TFs whose perturbation can restore gene expressions in a source state to target levels. Its effectiveness is demonstrated on datasets from yeast TF knockouts, cardiomyocytes from hypoplastic left heart syndrome patients, and mouse models of neurodegeneration. The method accurately identifies knocked-out TFs in the yeast dataset. In the cardiomyocyte dataset, it pinpoints TFs that, though not differentially expressed in many cases, exert significant regulatory influence on downstream differentially expressed genes. Finally, in the mouse model dataset, it identifies disease stage-specific TFs, improving similarity between healthy and diseased states at various time points. Unlike traditional approaches relying on differential expression analysis, our method uses network-based prioritization for more targeted and biologically relevant TF selection. These findings highlight its potential as a therapeutic tool for precise TF targeting to normalize gene expressions in diseased states.

bioinformatics↗

bioRxiv2024

Involvement of Hepatitis B Core-Related Antigen in Viral Genome Integration in Patients With Prior Hepatitis B Virus Infection

BackgroundThe incidence of non-B non-C hepatocellular carcinoma (NBNC-HCC), which is negative for hepatitis B surface antigen and hepatitis C virus antibodies, is on the rise. Relatively high numbers of NBNC-HCC patients are hepatitis B core antibody (HBcAb) positive, suggesting that previous HBV infection may play a role in NBNC-HCC development, though the exact mechanisms are unclear. This study aimed to investigate whether HBV genomes are integrated into the host genome of HBcAb-positive NBNC-HCC cases and how these integrations may contribute to cancer development and progression. MethodsHBV detection PCR using HBV-specific primers on DNA extracted from HBcAb-positive NBNC-HCC tissue samples was performed. Positive samples were further examined for HBV integration sites using viral DNA-capture sequencing. Additionally, hepatitis B core-related antigen (HBcrAg) serum levels were measured to assess whether they could be predictive for HBV detection PCR results. ResultsAmong 90 HBcAb-positive NBNC-HCC samples, HBV genome amplification was detected in 18 samples, and elevated HBcrAg levels were associated with the HBV detection PCR results. Seventeen of these samples exhibited HBV integration. The HBV genome was integrated near the TERT gene in 7 samples, resulting in significantly increased TERT mRNA levels; in the KMT2B gene (2 samples); and downstream of LOC441666 (2 samples). ConclusionThe integration sites we identified in our samples have been previously reported in HBV-related HCC, suggesting that HBV integration may also contribute to hepatocarcinogenesis in HBcAb-positive NBNC-HCC. Furthermore, HBcrAg could serve as a potential, noninvasive marker for detecting HBV integration in these cases.

microbiology↗

bioRxiv2024

Unraveling SARS-CoV-2 Host-Response Heterogeneity through Longitudinal Molecular Subtyping

Hospitalized COVID-19 patients exhibit diverse immune responses during acute infection, which are associated with a wide range of clinical outcomes. However, understanding these immune heterogeneities and their links to various clinical complications, especially long COVID, remains a challenge. In this study, we performed unsupervised subtyping of longitudinal multi-omics immunophenotyping in over 1,000 hospitalized patients, identifying two critical subtypes linked to mortality or mechanical ventilation with prolonged hospital stay and three severe subtypes associated with timely acute recovery. We confirmed that unresolved systemic inflammation and T-cell dysfunctions were hallmarks of increased severity and further distinguished patients with similar acute respiratory severity by their distinct immune profiles, which correlated with differences in demographic and clinical complications. Notably, one critical subtype (SubF) was uniquely characterized by early excessive inflammation, insufficient anticoagulation, and fatty acid dysregulation, alongside higher incidences of hematologic, cardiac, and renal complications, and an elevated risk of long COVID. Among the severe subtypes, significant differences in viral clearance and early antiviral responses were observed, with one subtype (SubC) showing strong early T-cell cytotoxicity but a poor humoral response, slower viral clearance, and greater risks of chronic organ dysfunction and long COVID. These findings provide crucial insights into the complex and context-dependent nature of COVID-19 immune responses, highlighting the importance of personalized therapeutic strategies to improve both acute and long-term outcomes.

immunology↗

bioRxiv2024

Syntopic call diversity among morphologically similar calling morphs ofthe paleotropical katydid of genus Mecopoda (Tettigoniidae) from an Indo-Burma biodiversity hotspot.

The acoustically diverse family of Tettigoniidae (Ensifera, Orthoptera) (commonly called bushcrickets or katydids), constitutes an excellent system to study the evolution of calling songs. Mecopoda is a genus that is particularly interesting because of the many morphologically cryptic calling morphs that have been described across South, East, and Southeast Asia, some of which are considered endemic to India. We describe five new syntopic and sympatric calling morphs of the genus Mecopoda that are morphologically similar but acoustically diverse from a sub-tropical forest in Meghalaya, in the Indo-Burma biodiversity hotspot. We report the morphological and acoustic characterization of these five call types, finding morphological overlap but well-separated acoustic clusters based on a PCA analysis of call temporal and spectral features. We also sequenced the COI gene mitochondrial gene of these five call types as well as other call types previously reported from India and performed a phylogenetic analysis of this gene relative to previously reported COI gene sequences from genus Mecopoda. Phylogenetic results suggest our five call types are not mutually monophyletic and relate from different subspecies of Mecopoda, of which three call types are genetically well separated from other calling morphs. This broadens our understanding of the evolution of acoustic diversity among paleotropical katydids.

zoology↗

bioRxiv2024

Reference-free Analysis of scRNA-seq Data Reveals Elevated rRNA and mtRNA Transcription during Neurogenesis in Axolotl

Analysis of single-cell RNA-seq data is typically performed on a gene expression matrix estimated by aligning reads to a reference transcriptome. However, this approach is difficult to apply to organisms with no or incomplete reference transcriptomes. In addition, events deviating from the reference remain undetected. Here we present a reference-free method to analyze single-cell RNA-seq data based on k-mers. We assess the performance of our method on a metastatic renal cell carcinoma dataset and find that it is largely able to capture differentially expressed genes. We then analyze a recently generated dataset to study neurogenesis in Axolotl and observe increased levels of transcription of rRNA and mtRNA during neurogenesis as well as a miRNA with previously predicted links to neuronal development. We also detect lncRNAs and intron retention in heart disease-related genes in diseased cardiomyocytes in an analysis of a congenital heart disease dataset.

bioinformatics↗

bioRxiv2024

RNA-DCGen: Dual Constrained RNA Sequence Generation with LLM-Attack

Designing RNA sequences with specific properties is critical for developing personalized medications and therapeutics. While recent diffusion and flow-matching-based generative models have made strides in conditional sequence design, they face two key limitations: specialization for fixed constraint types, such as tertiary structures, and lack of flexibility in imposing additional conditions beyond the primary property of interest. To address these challenges, we introduce RNA-DCGen, a generalized framework for RNA sequence generation that is adaptable to any structural or functional properties through straightforward finetuning with an RNA language model (RNA-LM). Additionally, RNA-DCGen can enforce conditions on the generated sequences by fixing specific conserved regions. On RNA generation conditioned on RNA distance maps, RNA-DCGen generates sequences with an average R2 score of 0.625 compared to random sequences that score only 0.118 over 250 generations as judged by a separate more capable RNA-LM. When conditioned on RNA secondary structures, RNA-DCGen achieves an average F1 score of 0.4 against a random baseline of 0.006.

bioinformatics↗

bioRxiv2024

Spatiotemporal dynamics of tumor-CAR T-cell interaction following local administration in solid cancers

The success of chimeric antigen receptor (CAR) T-cell therapy in treating hematologic malignancies has generated widespread interest in translating this technology to solid cancers. However, issues like tumor infiltration, the immunosuppressive tumor microenvironment, and tumor heterogeneity limit its efficacy in the solid tumor setting. Recent experimental and clinical studies propose local administration directly into the tumor or at the tumor site to increase CAR T-cell infiltration and improve treatment outcomes. Characteristics of the types of solid tumors that may be the most receptive to this treatment approach remain unclear. In this work, we develop a spatiotemporal model for CAR T-cell treatment of solid tumors, and use numerical simulations to compare the effect of introducing CAR T cells via intratumoral injection versus intracavitary administration in diverse cancer types. We demonstrate that the model can recapitulate tumor and CAR T-cell data from imaging studies of local administration of CAR T cells in mouse models. Our results suggest that locally administered CAR T cells will be most successful against slowly proliferating, highly diffusive tumors, which have the lowest average tumor cell density. These findings affirm the clinical observation that CAR T cells will not perform equally across different types of solid tumors, and suggest that measuring tumor density may be helpful when considering the feasibility of CAR T-cell therapy and planning dosages for a particular patient. We additionally find that local delivery of CAR T cells can result in deep tumor responses, provided that the initial CAR T-cell dose does not contain a significant fraction of exhausted cells.

cancer biology↗

bioRxiv2024

Interplay of Anchor and Latch Residues at Enzyme Active Site in Preserving Induced Fit Configurations for Efficient Catalysis: A Case Study of CARNMT1 for Histidine Methylation

Determining the origin of high catalytic efficiency of enzymes has been a central goal in biochemical research. But our understanding of the mechanisms by which enzymes achieve enormous rate enhancements is still incomplete. One of wildly accepted strategies to achieve rate enhancements by enzymes is to position catalytic residues precisely in enzyme-substrate complexes and form reactive conformations for catalysis. Such catalytic strategy through positioning may also serve as the basis for transition state stabilization. Although considerable efforts have been made to determine relationships between enzyme activities and the formation of reactive conformations of enzyme-substrate complexes, there is considerable uncertainty concerning the ability of enzymes to preserve reactive conformations during enzyme-catalyzed reactions. The failure of properly maintaining reactive conformations during reactions may prevent residues from exerting catalytic effects and lead to significant reductions of activities. Here QM/MM MD and free energy simulations are performed for CARNMT1 to determine the importance and requirements for preservation of catalytic competent reactive conformations during enzyme-catalyzed reactions. For CARNMT1, the existence of wild-type reactive conformation is directly related to the conformation of His347. Therefore, the conformation of His347 can be used to monitor preservation or collapse of the reactive conformation during catalysis. It is found that for wild-type the reactive conformation can not only be formed in the enzyme-substrate complex but also be preserved well after the reaction passes TS. For the Tyr386Ala, Tyr396Asp and Tyr398Ala mutants, the wild-type like reactive conformations can be formed in the mutant-substrate complexes but collapse in the early stage of reaction. Such pre-mature collapse of the reactive conformation in the mutants prevents the residues to fully exert catalytic effects and leads to a significant reduction of catalytic efficiency. The results suggest that the ability of enzymes to preserve reactive conformations with well-positioned catalytic residues during enzyme-catalyzed reactions can be an important factor for efficient catalysis, and this property of enzymes deserves attention.

biochemistry↗

bioRxiv2024

Actin isovariant ACT2-mediated cellular auxin homeostasis regulates lateral root organogenesis in Arabidopsis thaliana

Lateral root (LR) organogenesis is regulated by cellular flux of auxin within pericycle cells, which depends on the membrane distribution and polar localization of auxin carrier proteins. The correct distribution of auxin carrier proteins relies on the intracellular trafficking of these proteins aided by filamentous actin as a track. However, the precise role of actin in lateral root development is still elusive. Here, using vegetative class actin isovariant mutants, we revealed that loss of actin isovariant ACT8 led to an increase in lateral root formation. The distribution of auxin within pericycle cells was altered in act8 mutant, primarily due to the altered distribution of AUX1 and PIN7. Interestingly, incorporation of act2 mutant in act8 background (act2act8) effectively nullified the LR phenotype observed in act8 mutant, indicating that ACT2 plays an important role in LR development. To explore further, we investigated the possibility that the act8 mutants LR phenotype and cellular auxin distribution resulted from ACT2 overexpression. Consistent with the idea, enhanced lateral root formation, altered AUX1, PIN7 expression and auxin distribution in pericycle cells were observed in ACT2 overexpression lines. Collectively, these results suggest that actin isovariant ACT2 but not ACT8 plays a pivotal role in regulating source to sink auxin distribution during lateral root organogenesis.

developmental biology↗

bioRxiv2024

Beav: A bacterial genome and mobile element annotation pipeline

Comprehensive and accurate genome annotation is crucial for inferring the predicted functions of an organism. Numerous tools exist to annotate genes, gene clusters, mobile genetic elements, and other diverse features. However, these tools and pipelines can be difficult to install and run, be specialized for a particular element or feature, or lack annotations for larger elements that provide important genomic context. Integrating results across analyses is also important for understanding gene function. To address these challenges, we present the Beav annotation pipeline. Beav is a command-line tool that automates the annotation of bacterial genome sequences, mobile genetic elements, molecular systems and gene clusters, key regulatory features, and other elements. Beav uses existing tools in addition to custom models, scripts, and databases to annotate diverse elements, systems, and sequence features. Custom databases for plant-associated microbes are incorporated to improve annotation of key virulence and symbiosis genes in agriculturally important pathogens and mutualists. Beav includes an optional Agrobacterium-specific pipeline that identifies and classifies oncogenic plasmids and annotates plasmid-specific features. Following the completion of all analyses, annotations are consolidated to produce a single comprehensive output. Finally, Beav generates publication-quality genome and plasmid maps. Beav is on Bioconda and is available for download at https://github.com/weisberglab/beav.

bioinformatics↗

bioRxiv2024

Artemisinin-resistant Plasmodium falciparum Kelch13 mutant proteins display reduced heme-binding affinity and decreased artemisinin activation

The rapid emergence of artemisinin resistance (ART-R) poses a challenge to global malaria control efforts. ART potency is triggered by ferrous iron- and/or heme-mediated cleavage of the endoperoxide bond to generate reactive heme-ART alkoxy radicals and covalent heme-ART adducts that alkylate parasite targets or inhibit the detoxification of heme into {beta}-hematin crystals; both of which lead to parasite death. Mutations in the P. falciparum Kelch-containing protein Kelch13 (PfKekch13) confer clinical ART-R, in which the resistant parasites exhibit impaired hemoglobin uptake, reduced heme yield, and thus decreased ART activation. However, a more direct involvement of PfKelch13 in heme-mediated ART activation has not been reported. Here, we show that recombinant, purified PfKelch13 wild-type (WT) protein displays measurable binding affinity for both iron and heme, the main effectors for ART activation. Comparative biochemical analyses further indicate weaker heme-binding affinities in the two Southeast Asian ART-R PfKelch13 mutants C580Y and R539T compared to the ART-sensitive WT and A578S mutant proteins, which ultimately translates into reduced yield of heme-ART derivatives. In conclusion, this study provides the first evidence for regulated ART activation via the heme-binding propensity of PfKelch13, which may contribute towards modulating the level of ART-R in malaria parasites with PfKelch13 mutations.

biochemistry↗

bioRxiv2023

Probabilistic clustering of cells using single-cell RNA-seq data

Single-cell RNA sequencing is a modern technology for analyzing cellular heterogeneity. A key challenge is to cluster a heterogeneous sample of different cell types into multiple different homogeneous groups. Although there exist a number of clustering methods, they do not perform well consistently across various datasets. Moreover, most of them are not based on probabilistic approaches making it difficult to assess uncertainties in their results. Therefore, in spite of having large cell atlases, it is often quite difficult to map cells to types. In addition, many of the methods require prior knowledge such as marker gene information for each type. Also due to technological limitations, dropouts of gene expressions may occur in the data which is not taken into account in other methods. Here we present a probabilistic method named CellHorizon for clustering scRNA-seq data that is based on a generative model, handles dropouts and works without any prior marker gene information. Experiments reveal that our method outperforms current state-of-the-art methods overall on six gold standard datasets.

bioinformatics↗

bioRxiv2023

Alignment Free Phylogeny Construction Using Maximum Likelihood Using k-mer Counts

Estimating phylogenetic trees from molecular data often involves first performing a multiple sequence alignment of the sequences and then identifying the tree that maximizes likelihood computed under a model of nucleotide substitution. However, sequence alignment is computationally challenging for long sequences, especially in the presence of genomic rearrangements. To address this, methods for constructing phylogenetic trees without aligning the sequences i.e. alignment-free methods have been proposed. They are generally fast and can be used to construct phylogenetic trees of a large number of species but they primarily estimate phylogenies by computing pairwise distances and are not based on statistical models of molecular evolution. In this paper, we introduce a model for k-mer frequency change based on a birth-death-migration process which can be used to estimate maximum likelihood phylogenies from frequencies of k-mers in genomic sequences of species in an alignment-free approach. Experiments on real and simulated data demonstrate the efficacy of the model for likelihood based alignment-free phylogeny construction.

evolutionary biology↗

bioRxiv2023

Early life adversity has sex-dependent effects on survival across the lifespan in rhesus macaques

Exposure to adversity during early life is linked to lasting detrimental effects on evolutionary fitness across many taxa. However, due to the challenges of collecting longitudinal data, especially in species where one sex disperses, direct evidence from long-lived species remains relatively scarce. Here we test the effects of early life adversity on male and female longevity in a free-ranging population of rhesus macaques (Macaca mulatta) at Cayo Santiago, Puerto Rico. We leveraged six decades of data to quantify the relative importance of ten forms of early life adversity for 6,599 macaques (3,230 male, 3,369 female), with a smaller sample size (N=299) for one form of adversity (maternal social isolation) which required high-resolution behavioral data. We found that individuals who experienced more early life adversity died earlier than those who experienced less adversity. Mortality risk was highest during early life, defined as birth to four years old, suggesting acute survival effects of adversity, but heightened mortality risk was also present in macaques who survived to adulthood. Females and males were affected differently by some forms of adversity, and these differences might be driven by varying energetic demands, female philopatry, and male dispersal. By leveraging data on thousands of macaques collected over decades, our results show that the fitness consequences of early life adversity are not uniform across individuals but vary as a function of the type of adversity, timing, and social context, and thus contribute to our limited but growing understanding of the evolution of early life sensitivities in long-lived species. Significance Statement Exposure to early life adversity, even when conditions subsequently improve, can have profound and persistent consequences for human health. Negative effects of early life adversity appear widespread across the animal kingdom. To date, however, direct evidence from long-lived species is relatively scarce due to the difficulties of collecting data from early life till death. We leverage six decades of observations on thousands of free-ranging male and female rhesus macaques to examine the complex ways early life adversity impacts survival. Our results suggest that the type of adversity and life history factors intersect to impact immediate and downstream survival. By studying early life adversity across environments, cultures, contexts, and species, we can better understand the evolutionary underpinnings of early life sensitivities.

animal behavior and cognition↗

bioRxiv2023

HIV-Tocky system to visualize proviral expression dynamics

The stably integrated pool of HIV-1 proviruses in the host genome stands against curative strategies. This reservoir is extremely heterogeneous with respect to host cell type, anatomical location, integration site, and replication fitness. During the initial phase of infection, only a few infected cells can resist host immune clearance or cytopathic effect and establish this resistant pool. The mechanisms underlying HIV latency initiation are not fully resolved yet. In the current study, we propose and validate a new reporter model for monitoring HIV-1 provirus silencing and reactivation using Timer of cell kinetics and activity (Tocky). HIV-Tocky system uses a fluorescent Timer protein whose emission spectrum spontaneously shifts from blue to red to reveal HIV-1 provirus dynamics. We dissected provirus transcriptional phases into early, persistent, recently silenced, and latent. To our knowledge, this is the first report to distinguish two latent subsets: a directly non-expressing population and a recently silenced after brief expression. In-depth integration site analysis suggested that the distribution of proviruses in directly latent cells was similar to that in actively transcribing cell population, whereas recently silenced cells tended to harbor proviruses integrated into heterochromatin. Furthermore, we established a library of various single integration clones at which we utilized to demonstrate the efficiency of the block-and-lock strategy by capturing the fast dynamics of silencing that were overlooked in previous models. In summary, we propose HIV-Tocky system to serve as a time-sensitive model that can capture the dynamics of provirus expression, making it a useful tool for HIV latency research. Significance StatementDeterminants of HIV-1 latency establishment are yet to be elucidated. This reservoir comprises a rare fraction of infected cells that can survive host and virus-mediated killing. In vitro reporter models so far offered a feasible means to inspect this population, but with limited capabilities to dissect provirus silencing dynamics. Here, we describe a new HIV reporter model (HIV-Tocky) with dual fluorescence spontaneous shifting to reveal provirus silencing and reactivation dynamics. This unique feature allowed; for the first time, identifying two latent populations: a directly latent, and a recently silenced subset, with the latter having integration features suggestive of stable latency. Our proposed model can help address the heterogeneous nature of HIV reservoirs and offers new possibilities for evaluating eradication strategies. ClassificationBiological Sciences, Microbiology.

microbiology↗

bioRxiv2023

Determining the shelf life of an erythropoietin alfa biosimilar GBPD002 through stability study

GBPD002 is a recombinant human erythropoietin (rhEPO) produced by recombinant DNA technology using mammalian cell expression system. In this study, samples were analyzed according to current Good Laboratory Practice (cGLP) and regulatory guidelines to evaluate the quality of the product under the influence of variety of environmental factors in a time-dependent manner. Accelerated (25 {+/-} 2 {degrees}C and relative humidity: 60 {+/-} 5 %) and real-time (5 {+/-} 3 {degrees}C) stability study were conducted up to 6 months and 12 months, respectively; samples were analyzed in every 3 months. After 12 months, real-time stability studies were performed with 6 months interval up to18 months, which to be continued up to 24 months. Appearance and pH were assessed using standard methods, and molecular weight was determined by Western blotting. Chromatographic identification and quantitative assays were performed by reversed- phase chromatography (RPC). High-molecular weight aggregatesand degradants were determined using size exclusion chromatography (SEC) and particle size distribution (PSD) analysis. Biofunctionality of the samples were evaluated by in vitro and in vivo bioassays. Bacterial endotoxin and sterility test were performed as safety parameters. All samples met the acceptance criteria, and the data were extrapolated using the regulatory guideline to determine the shelf life. The data revealed that the GBPD002 is stable up to 24 months.

pharmacology and toxicology↗

bioRxiv2023

Indigenous probiotics Lactobacillus reuteri and Enterococcus faecium exhibit positive growth performance and disease prevention against extended-spectrum cephalosporin and fluoroquinolones resistant Salmonella enterica in broiler chicks.

ABSTRACTThe rapid increase in antibiotic resistance poses a global threat to public health, necessitating the development of effective antimicrobial alternatives. This study compared an indigenous probiotic mix containing Lactobacillus reuteri and two strains of Enterococcus faecium to a commercial probiotic blend ProtexinR on the growth performance, mortality rate, histomorphology, serum immunoglobulins, and intestinal microflora of broiler chickens challenged with two multi drug resistant Salmonella serovars, Typhimurium and Enteritidis. Two hundred and forty day-old broiler chicks were randomly assigned to six treatment groups for 4 weeks: the treatment groups were; birds continuously supplemented with only indigenous probiotic strains (108 CFU/mL) (IPRO-); birds challenged with Salmonella serovars 106 (CFU/mL) (PC+); birds continuously supplemented with indigenous probiotic strains and challenged with Salmonella serovars (IPRO+); birds supplemented with ProtexinR and challenged with Salmonella serovars (CM+); birds supplemented with only ProtexinR (CM-); and birds with no Salmonella challenge or probiotics (negative control; PC-). The results revealed that IPRO- diets significantly improved feed conversion ratio (FCR) and increased body weight (BW) (P [≤] 0.05). No effect of probiotic treatments was observed on IPRO- and CM- on relative organ weights as compared to the negative control (PC-). The Salmonella- challenged group PC+ had the highest (20%) mortality rate and lowest BW. The IPRO- had significantly lower FCR (1.55) compared to PC- (1.86) and PC+ (1.95). The broilers in the IPRO- group showed significantly increased serum concentrations of IgA and IgG relative to both control groups (P [≤] 0.05). Morphological analysis of the ileum revealed significant increases (P [≤] 0.05) in the villus height and villus height/crypt depth in birds fed IPRO- compared with the PC+. Cecal Lactobacillus and Enterococcus counts were the highest (P [≤] 0.05) and Salmonella counts were the lowest (P [≤] 0.05) in the IPRO- group compared to the Salmonella infected group PC+. These results indicated that indigenous probiotic strains Lactobacillus reuteri and Enterococcus faecium can be an effective and low-cost alternative compared to commercial probiotics in the Pakistan poultry industry.

microbiology↗