A protocol for generating germ-free Heligmosomoides polygyrus bakeri larvae for gnotobiotic helminth infection studies

The influence of the protein component in the diet of the host on the population dynamics of gastrointestinal helminth infection was studied by using a mouse – H. polygyrus experimental model. Mice fed a 2% (by mass) protein diet ad libitum maintained body weight during the experiment, but gained weight steadily when fed a diet containing 8% (by mass) protein. When repeatedly infected with 5, 10, 20 or 40 larvae every 2 weeks, the mice fed the 2% (by mass) protein diet accumulated adult worms in direct proportion to exposure to the infective stages. Under similar infection régimes, mice fed an 8% (by mass) protein diet acquired a partly effective immunity to reinfection by the nematode. Acquired immunity was principally manifest as a reduction in the survival of adult worms, although a slight increase in the mortality rate and/or the development time of the tissue-dwelling larval phase was observed. Worm fecundity per head was significantly depressed in hosts fed the 8% protein diet. In conclusion, in these experiments it is demonstrated that the nutritional status of the host can influence the population dynamics of helminth infection.

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The gut microbiota limits Th2 immunity to Heligmosomoides polygyrus bakeri infection

10.1101/2020.01.30.927111 ◽

2020 ◽

Author(s):

Gabriel A. Russell ◽

Cynthia Faubert ◽

Elena F. Verdu ◽

Irah L. King

Keyword(s):

Gut Microbiota ◽

Culture Method ◽

Th2 Response ◽

Heligmosomoides Polygyrus ◽

Germ Free ◽

Commensal Microbiota ◽

Mutualistic Relationship ◽

Th2 Immunity

ABSTRACTHelminth-induced alterations to the gut microbiota have been shown to affect immune responses at local and peripheral sites. Studies examining helminth-microbiota interactions, however, have been limited due to the practical constraints of performing germ-free experiments with parasites that thrive in microbial-rich conditions to complete their development. The infectious (L3) larvae of the murine helminth Heligmosomoides polygyrus bakeri (Hpb), for example, are normally reared using a fecal-culture method and therefore are inherently unsuitable for germ-free studies in vivo. Herein, we detail an adapted methodology for rearing effectively germ-free Hpb larvae that are able to maintain the axenic status of a germ-free host during infection. We validate that these larvae do not display any fitness defects relative to fecal-grown larvae and evoke a comparable immune response in vivo. Characterization of axenic Hpb infection reveals that the commensal microbiota play a multifaceted role during infection - curbing the anti-Hpb Th2 response and directing the resolution of tissue granulomas, while simultaneously promoting parasite fitness. Overall these data demonstrate a mutualistic relationship between commensal microbes, enteric helminths and the infected host.

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Inhibition of miR-99a-5p prevents allergen-driven airway exacerbations without compromising type-2 memory responses in the intestine following helminth infection

Mucosal Immunology ◽

10.1038/s41385-021-00401-x ◽

2021 ◽

Author(s):

Lewis Entwistle ◽

Helena Aegerter ◽

Stephanie Czieso ◽

Eleni Amaniti ◽

Riccardo Guidi ◽

...

Keyword(s):

Molecular Mechanisms ◽

Helminth Infection ◽

Smooth Muscle Contraction ◽

Therapeutic Interventions ◽

Transcriptional Analysis ◽

Intestinal Helminth ◽

Heligmosomoides Polygyrus ◽

Minimal Impact ◽

Acute Exacerbations

AbstractAcute exacerbations (AE) of asthma, remain one of the biggest concerns for patients living with asthma. As such, identifying the causes, the molecular mechanisms involved and new therapeutic interventions to prevent AE is a high priority. Immunity to intestinal helminths involves the reactivation of type-2 immune responses leading to smooth muscle contraction and mucus hypersecretion–physiological processes very similar to acute exacerbations in the airways following allergen exposure. In this study, we employed a murine model of intestinal helminth infection, using Heligmosomoides polygyrus, to identify miRNAs during active expulsion, as a system for the identification of miRNAs that may contribute to AE in the airways. Concomitant with type-2 immunity and expulsion of H. polygyrus, we identified miR-99a-5p, miR-148a-3p and miR-155-5p that were differentially regulated. Systemic inhibition of these miRNAs, alone or in combination, had minimal impact on expulsion of H. polygyrus, but inhibition of miR-99a-5p or miR-155-5p significantly reduced house dust mite (HDM)-driven acute inflammation, modelling human acute exacerbations. Immunological, pathological and transcriptional analysis identified that miR-155-5p or miR-99a-5p contribute significantly to HDM-driven AE and that transient inhibition of these miRNAs may provide relief from allergen-driven AE, without compromising anti-helminth immunity in the gut.

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Infection with a small intestinal helminth Heligmosomoides polygyrus bakeri consistently alters microbial communities throughout the small and large intestine

10.1101/575787 ◽

2019 ◽

Author(s):

Alexis Rapin ◽

Audrey Chuat ◽

Luc Lebon ◽

Mario M. Zaiss ◽

Benjamin Marsland ◽

...

Keyword(s):

Microbial Communities ◽

Large Intestine ◽

Helminth Infection ◽

Entire Length ◽

Intestinal Helminth ◽

Mammalian Host ◽

Heligmosomoides Polygyrus ◽

Small Intestinal ◽

The Impact ◽

Small And Large Intestine

AbstractIncreasing evidence suggests that intestinal helminth infection can alter intestinal microbial communities with important impacts on the mammalian host. However, all of the studies to date utilize different techniques to study the microbiome and access different sites of the intestine with little consistency noted between studies. In the present study, we set out to perform a comprehensive analysis of the impact of intestinal helminth infection on the mammalian intestinal bacterial microbiome. For this purpose, we investigated the impact of experimental infection using the natural murine small intestinal helminth,Heligmosomoides polygyrus bakeri(Hpb) and examined possible alterations in both the mucous and luminal bacterial communities along the entire small and large intestine. We also explored the impact of common experimental variables, including the parasite batch and pre-infection microbiome, on the outcome of helminth-bacterial interactions. This work provides evidence that helminth infection reproducibly alters intestinal microbial communities – with an impact of infection noted along the entire length of the intestine. Although the exact nature of helminth-induced alterations to the intestinal microbiome differed depending on the parasite batch and microbiome community structure present prior to infection, changes extended well beyond the introduction of new bacterial species by the infecting larvae. Moreover, striking similarities between different experiments were noted, including the consistent outgrowth of a bacterium belonging to the Peptostreptococcaceae family throughout the intestine.Author SummaryIncreasing evidence indicates a role for interactions between intestinal helminths and the microbiome in regulating mammalian health, and a greater understanding of helminth-microbiota interactions may open the path for the development of novel immunomodulatory therapies. However, such studies are hampered by the inconsistent nature of the data reported so far. Such inconsistancies likely result from variations in the experimental and technological methodologies employed to investigate helminth-microbiota interactions and well has natural variation in the starting microbiome composition and/or worm genetics. We conducted a thorough study in which the reproducibility of helminth-induced alterations of microbial communities was determined and impact of common experimental variables – such as the starting microbiome and parasite batch - was determined. Our work reveals the robust ability of small intestinal helminth infection to alter microbial communities along the entire length of the intestine and additionally identifies a single bacterium that is strongly associated with infection across multiple experiments.

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Small intestinal levels of the branched short-chain fatty acid isovalerate are elevated during infection with Heligmosomoides polygyrus and can promote helminth fecundity

Infection and Immunity ◽

10.1128/iai.00225-21 ◽

2021 ◽

Author(s):

Mia H. E. Kennedy ◽

Tara P. Brosschot ◽

Katherine M. Lawrence ◽

Rachael D. FitzPatrick ◽

Jenna M. Lane ◽

...

Keyword(s):

Small Intestine ◽

Short Chain Fatty Acid ◽

Helminth Infection ◽

Chain Fatty Acid ◽

Isovaleric Acid ◽

Heligmosomoides Polygyrus ◽

Bacterial Microbiota ◽

Proximal Small Intestine ◽

Small Intestinal ◽

Helminth Fecundity

Heligmosomoides polygyrus is a helminth which naturally infects mice and is widely used as a laboratory model of chronic small intestinal helminth infection. While it is known that infection with H. polygyrus alters the composition of the host’s bacterial microbiota, the functional implications of this alteration are unclear. We investigated the impact of H. polygyrus infection on short-chain fatty acid (SCFA) levels in the mouse intestine and sera. We found that helminth infection resulted in significantly upregulated levels of the branched SCFA isovaleric acid, exclusively in the proximal small intestine, which is the site of H. polygyrus colonization. We next set out to test the hypothesis that elevating local levels of isovaleric acid was a strategy used by H. polygyrus to promote its own fitness within the mammalian host. To test this, we supplemented the drinking water of mice with isovalerate during H. polygyrus infection and examined whether this affected helminth fecundity or chronicity. We did not find that isovaleric acid supplementation affected helminth chronicity, however, we found that it did promote helminth fecundity, as measured by helminth egg output in the feces of mice. Through antibiotic-treatment of helminth-infected mice, we found that the bacterial microbiota was required in order to support elevated levels of isovaleric acid in the proximal small intestine during helminth infection. Overall, our data reveal that during H. polygyrus infection there is a microbiota-dependent localized increase in the production of isovaleric acid in the proximal small intestine and this supports helminth fecundity in the murine host.

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Anti-Inflammatory Mechanisms of Enteric Heligmosomoides polygyrus Infection against Trinitrobenzene Sulfonic Acid-Induced Colitis in a Murine Model

Infection and Immunity ◽

10.1128/iai.00744-07 ◽

2008 ◽

Vol 76 (10) ◽

pp. 4772-4782 ◽

Cited By ~ 50

Author(s):

Thomas L. Sutton ◽

Aiping Zhao ◽

Kathleen B. Madden ◽

Justin E. Elfrey ◽

Blaine A. Tuft ◽

...

Keyword(s):

Inflammatory Bowel Disease ◽

Mast Cell ◽

Sulfonic Acid ◽

Helminth Infection ◽

Cytokine Expression ◽

Trinitrobenzene Sulfonic Acid ◽

Heligmosomoides Polygyrus ◽

Th2 Cytokine ◽

Inflammatory Bowel ◽

Th1 Cytokines

ABSTRACT Recent studies showed that enteric helminth infection improved symptoms in patients with inflammatory bowel disease as well as in experimental models of colitis. The aim of this study was to determine the mechanism of the protective effect of helminth infection on colitis-induced changes in immune and epithelial cell function. BALB/c mice received an oral infection of Heligmosomoides polygyrus third-stage larvae, were given intrarectal saline or trinitrobenzene sulfonic acid (TNBS) on day 10 postinfection, and were studied 4 days later. Separate groups of mice received intrarectal saline or TNBS on day 10 and were studied on day 14. Muscle-free colonic mucosae were mounted in Ussing chambers to measure mucosal permeability and secretion. Expression of cytokines was assessed by quantitative real-time PCR, and mast cells were visualized by immunohistochemistry. TNBS-induced colitis induced mucosal damage, upregulated Th1 cytokines, and depressed secretory responses. Heligmosomoides polygyrus elevated Th2 cytokine expression, increased mast cell infiltration and mucosal resistance, and also reduced some secretory responses. Prior H. polygyrus infection prevented TNBS-induced upregulation of Th1 cytokines and normalized secretory responses to specific agonists. TNBS-induced colitis did not alter H. polygyrus-induced mast cell infiltration or upregulation of Th2 cytokine expression. The results indicate that the protective mechanism of enteric nematode infection against TNBS-induced colitis involves prevention of Th1 cytokine expression and improved colonic function by a mechanism that may involve mast cell-mediated protection of neural control of secretory function. Similar response patterns could account for the clinical improvement seen in inflammatory bowel disease with helminthic therapy.

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Helminth Infection Can Reduce Insulitis and Type 1 Diabetes through CD25- and IL-10-Independent Mechanisms

Infection and Immunity ◽

10.1128/iai.01170-08 ◽

2009 ◽

Vol 77 (12) ◽

pp. 5347-5358 ◽

Cited By ~ 87

Author(s):

Qian Liu ◽

Krishnan Sundar ◽

Pankaj K. Mishra ◽

Gity Mousavi ◽

Zhugong Liu ◽

...

Keyword(s):

T Cells ◽

Type 1 Diabetes ◽

Helminth Infection ◽

Inflammatory Responses ◽

Nod Mice ◽

Interleukin 4 ◽

Heligmosomoides Polygyrus ◽

Nonobese Diabetic

ABSTRACT Parasitic helminth infection has been shown to modulate pathological inflammatory responses in allergy and autoimmune disease. The aim of this study was to examine the effects of infection with a helminth parasite, Heligmosomoides polygyrus, on type 1 diabetes (T1D) in nonobese diabetic (NOD) mice and to elucidate the mechanisms involved in this protection. H. polygyrus inoculation at 5 weeks of age protected NOD mice from T1D until 40 weeks of age and also inhibited the more aggressive cyclophosphamide-induced T1D. Moreover, H. polygyrus inoculation as late as 12 weeks of age reduced the onset of T1D in NOD mice. Following H. polygyrus inoculation of NOD mice, pancreatic insulitis was markedly inhibited. Interleukin-4 (IL-4), IL-10, and IL-13 expression and the frequency of CD4+ CD25+ FoxP3+ regulatory T cells were elevated in mesenteric and pancreatic lymph nodes. Depletion of CD4+ CD25+ T cells in vivo did not abrogate H. polygyrus-induced T1D protection, nor did anti-IL-10 receptor blocking antibody. These findings suggest that infection with H. polygyrus significantly inhibits T1D in NOD mice through CD25- and IL-10-independent mechanisms and also reduces the severity of T1D when administered late after the onset of insulitis.

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Intestinal helminth infection enhances bacteria-induced recruitment of neutrophils to the airspace

Scientific Reports ◽

10.1038/s41598-019-51991-3 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Shao Rong Long ◽

Bernard B. Lanter ◽

Michael A. Pazos ◽

Hongmei Mou ◽

Juliana Barrios ◽

...

Keyword(s):

Structural Changes ◽

Helminth Infection ◽

Neutrophil Infiltration ◽

Intestinal Helminth ◽

Cell Hyperplasia ◽

Heligmosomoides Polygyrus ◽

Host Immune Responses ◽

Th2 Cytokine ◽

Helminth Infections

Abstract Intestinal helminth infections elicit Th2-type immunity, which influences host immune responses to additional threats, such as allergens, metabolic disease, and other pathogens. Th2 immunity involves a shift of the CD4+ T-cell population from type-0 to type-2 (Th2) with increased abundance of interleukin (IL)-4 and IL-13. This study sought to investigate if existing gut-restricted intestinal helminth infections impact bacterial-induced acute airway neutrophil recruitment. C57BL/6 mice were divided into four groups: uninfected; helminth-Heligmosomoides polygyrus infected; Pseudomonas aeruginosa infected; and coinfected. Mice infected with H. polygyrus were incubated for 2 weeks, followed by P. aeruginosa intranasal inoculation. Bronchial alveolar lavage, blood, and lung samples were analyzed. Interestingly, infection with gut-restricted helminths resulted in immunological and structural changes in the lung. These changes include increased lung CD4+ T cells, increased Th2 cytokine expression, and airway goblet cell hyperplasia. Furthermore, coinfected mice exhibited significantly more airspace neutrophil infiltration at 6 hours following P. aeruginosa infection and exhibited an improved rate of survival compared with bacterial infected alone. These results suggest that chronic helminth infection of the intestines can influence and enhance acute airway neutrophil responses to P. aeruginosa infection.

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Impaired host resistance to Salmonella during helminth co-infection is restored by anthelmintic treatment prior to bacterial challenge

PLoS Neglected Tropical Diseases ◽

10.1371/journal.pntd.0009052 ◽

2021 ◽

Vol 15 (1) ◽

pp. e0009052

Author(s):

Tara P. Brosschot ◽

Katherine M. Lawrence ◽

Brandon E. Moeller ◽

Mia H. E. Kennedy ◽

Rachael D. FitzPatrick ◽

...

Keyword(s):

Small Intestine ◽

Host Resistance ◽

Helminth Infection ◽

Bacterial Colonization ◽

Drug Clearance ◽

Anthelmintic Treatment ◽

Intestinal Lumen ◽

Intestinal Helminth ◽

Heligmosomoides Polygyrus ◽

Small Intestinal

Intestinal helminth infection can impair host resistance to co-infection with enteric bacterial pathogens. However, it is not known whether helminth drug-clearance can restore host resistance to bacterial infection. Using a mouse helminth-Salmonella co-infection system, we show that anthelmintic treatment prior to Salmonella challenge is sufficient to restore host resistance to Salmonella. The presence of the small intestine-dwelling helminth Heligmosomoides polygyrus at the point of Salmonella infection supports the initial establishment of Salmonella in the small intestinal lumen. Interestingly, if helminth drug-clearance is delayed until Salmonella has already established in the small intestinal lumen, anthelmintic treatment does not result in complete clearance of Salmonella. This suggests that while the presence of helminths supports initial Salmonella colonization, helminths are dispensable for Salmonella persistence in the host small intestine. These data contribute to the mechanistic understanding of how an ongoing or prior helminth infection can affect pathogenic bacterial colonization and persistence in the mammalian intestine.

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Staphylococcal bacteremia in a "Germ-free" unit

Archives of Internal Medicine ◽

10.1001/archinte.136.11.1238 ◽

1976 ◽

Vol 136 (11) ◽

pp. 1238-1240 ◽

Cited By ~ 1

Author(s):

M. E. Plaut

Keyword(s):

Germ Free

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