This study was supported by the Research Institute of the McGill University Health Centre (RI-MUHC), Canadian Institutes of Health Research (CIHR). This study was made possible by the following funding agencies, institutions, and foundations: Canadian Institute of Health Research (CIHR), Foundation of Stars, Fonds de Recherche Qu&#233;bec-Sciences (FRQS), McGill University, and Sherbrooke University. Many thanks to Dr. Claire Poyart, University Denis Diderot (Paris VII), France, and Dr. Mariela Segura, University de Montr&#233;al, Canada for the generous gifts of GBS.

All experiments were approved by the Research Institute of McGill University Health Centre (RI-MUHC). All experiments were performed according to the Canadian Council on Animal Care.
1. Pregnant Lewis rats
<ol>
	<li>Obtain Lewis rats from commercial sources at gestational day (G)14. House them in an appropriate animal facility (RI-MUHC animal facility) in a controlled environment at 20-23 &#176;C with a 12 h light/dark cycle, and access to water and food ad libitum<...

Critical steps in the protocol 
Several steps of the protocol are critical and require some quality controls. For instance, there is a risk of contamination of the GBS stock by other pathogens. This can be rapidly identified using the appropriate technique of GBS microbial identification such as colony aspect on BHI agar (e.g., size, shape, color), plating in duplicate the &#946;-hemolytic GBS dose on Columbia blood agar with 5% sheep blood medium and on CHROMID Strepto B agar, a selective chromogen...

Maternal immune activation (MIA) has been described as one of the most critical independent risk factors for premature birth, fetal death, and lifelong cognitive and behavioral impairments in the progeny1,2,3,4. Much of the existing preclinical research about the role of gestational inflammation on placental and developmental outcomes uses pathogen components, such as lipopolysaccharide (LPS) from E. coli and the synthetic analog of viral double-stranded RNA, polyinosinic: polycytidylic acid (Poly[I: C]), that mimic viral infections. However, even though Group B Streptococcus (GBS) is the most frequent cause of perinatal infection, few animal models have addressed its role in inflammatory mechanisms at play and the outcomes5.
GBS is an encapsulated gram-positive coccus that colonizes the lower genital tract in approximately 15%-30% of pregnant women6. It leads to placental infection/inflammation, termed chorioamnionitis7,8. Of the ten GBS serotypes, the two most frequent serotypes Ia and III are major infectious determinants of injuries in maternofetal tissues9,10.&#160;GBS infection has been shown to lead to a higher inflammatory response in fetal blood and placental deficiency, which are highly suspected to be involved in multiple neurodevelopmental disorders such as cerebral palsy (CP), attention deficit hyperactivity disorder (ADHD), and autism spectrum disorder (ASD)5,11.
Over the past ten years, we have developed a rat model of GBS-induced chorioamnionitis that leads to various developmental impairments in the offspring12. This preclinical model demonstrates the causal link between GBS-induced placental inflammation and a range of sex-specific neurodevelopmental impairments in the offspring13,14,15. The goal of this article is to provide readers with insight into the design of a preclinical rat model of end-gestational infection and resulting neuro-behavioral impairments in the offspring. The present protocol aims to mimic the clinical reality of GBS-induced chorioamnionitis.
Results from this preclinical model show that end-gestational intra-peritoneal (IP) inoculation (Figure 1) of GBS leads to (i) placental infection and inflammation, fulfilling the diagnostic criteria of chorioamnionitis16; (ii) a massive upregulation of IL-1&#946; and downstream inflammatory molecules from the IL-1-pathway, within the placenta12; (iii) neurodevelopmental impairments in the offspring12; (iv) sex differences in immune responses and subsequent neurobehavioral impairments, such as female offspring presenting adult ADHD-like traits while male offspring present early-onset and long-lasting ASD-like traits; (v) distinct neurobehavioral outcomes in the progeny depending on the GBS serotype used to induce chorioamnionitis14,15. In line with these findings, the main next steps utilizing this model will be to test, firstly, the role of androgen in GBS-induced chorioamnionitis and, secondly, the placental- and neuro-protective role of molecules targeting specific inflammatory pathways, in the hope to bring some of these molecules to the threshold of therapeutic clinical trials.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>5 mL sterile tube</td>
			<td>BD Biosciences</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>50 ml falcon tubes</td>
			<td>Thermo Fisher</td>
			<td>339652</td>
			<td></td>
		</tr>
		<tr>
			<td>Blade or scalpel</td>
			<td>BD Medical</td>
			<td>371716</td>
			<td></td>
		</tr>
		<tr>
			<td>Brain Heart Infusion Broth</td>
			<td>Criterion (Hardy diagnostics)</td>
			<td>C5141</td>
			<td></td>
		</tr>
		<tr>
			<td>CHROMID Strepto B agar plate</td>
			<td>BioMerieux, Saint-Laurent</td>
			<td>43461</td>
			<td></td>
		</tr>
		<tr>
			<td>Columbia blood agar 5 % with sheep blood medium</td>
			<td>Thermo Scientific</td>
			<td>R01215</td>
			<td></td>
		</tr>
		<tr>
			<td>Forward primer</td>
			<td>5&#39; - TAC AGC CTG AGG ACA TAT TA3&#39;</td>
			<td>Sigma</td>
			<td></td>
		</tr>
		<tr>
			<td>Insulin syringe</td>
			<td>Becton, Dickinson and Co(BD)</td>
			<td>324702</td>
			<td></td>
		</tr>
		<tr>
			<td>Lewis rats</td>
			<td>Charles River Laboratories</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Methylbutan</td>
			<td>Sigma Aldrich</td>
			<td>M32631</td>
			<td></td>
		</tr>
		<tr>
			<td>Microtainer blood collection tubes</td>
			<td>Becton, Dickinson and Co(BD)</td>
			<td>365965</td>
			<td></td>
		</tr>
		<tr>
			<td>Reverse primer</td>
			<td>5&#39; - GCA CTT TAA CCC TTC GAT GA -3&#39;</td>
			<td>Sigma</td>
			<td></td>
		</tr>
		<tr>
			<td>Serological Pipettes 1 ML</td>
			<td>Thermo Fisher</td>
			<td>170353N</td>
			<td></td>
		</tr>
		<tr>
			<td>Serological Pipettes 10 ML</td>
			<td>Thermo Fisher</td>
			<td>170356N</td>
			<td></td>
		</tr>
		<tr>
			<td>Serological Pipettes 25 ML</td>
			<td>Thermo Fisher</td>
			<td>170357N</td>
			<td></td>
		</tr>
		<tr>
			<td>Serological Pipettes 5 ML</td>
			<td>Thermo Fisher</td>
			<td>170355N</td>
			<td></td>
		</tr>
		<tr>
			<td>Superfrost Plus Micro Slide, Premium</td>
			<td>VWR</td>
			<td>CA48311-703</td>
			<td></td>
		</tr>
	</tbody>
</table>

c-section of preclinical animal model of chorioamnionitis triggered by group b streptococcus (gbs)

Group B Streptococcus (GBS) is one of the most common bacteria isolated during human pregnancy. It is a leading cause of placental infection/inflammation, termed chorioamnionitis. Chorioamnionitis exposes the developing fetus to a high risk of organ injuries, perinatal morbidity, and mortality, as well as life-long neurobehavioral impairments and other non-neurological developmental issues. The two most frequent subtypes of GBS isolates from maternal and fetal tissues are serotypes Ia (13%-23%) and III (25%-53%). Our lab has developed and characterized a rat model of GBS-induced chorioamnionitis to study subsequent impacts on the central nervous system of the developing fetus and to understand underlying mechanistic aspects. This article presents the design as well as uses of the preclinical rat model, which closely reproduces the hallmark of GBS-induced chorioamnionitis in humans. This article aims to help scientists reproduce the experimental design as well as to provide support through examples of troubleshooting. The present model may also contribute to potential discoveries through uncovering causes, mechanisms, and novel therapeutic avenues, which remain unsettled in many developmental impairments arising from chorioamnionitis. Furthermore, the use of this model may be extended to the studies of perinatal non-neurological common and severe morbidities affecting, for instance, the retina, bowel, lung, and kidney. The main interest of this research is in the field of GBS-induced fetal neurodevelopmental impairments such as cerebral palsy (CP), attention deficit hyperactivity disorder (ADHD), and autism spectrum disorder (ASD). The rationale supporting this model is presented in this article, followed by procedures and results.

IP inoculation of GBS resulted in placental infection 
Immunohistochemistry (IHC) (using polyclonal antibodies targeting GBS serotype Ia) staining showed that GBS infection reached the decidual compartment of the placenta. Infection also spread from the decidua to the labyrinth, chorionic plate, and in some instances, to fetuses leading to fetal death (5.8 &#177; 0.8 in GBS-exposed vs. 9.3 &#177; 0.6 pups in control (CTL) litters)18. Hence, the litter size was decreased at bi...

Read this Scientific Article Protocol about C-section of Preclinical Animal Model of Chorioamnionitis Triggered by Group B Streptococcus GBS at JoVE.com

C-section of Preclinical Animal Model of Chorioamnionitis Triggered by Group B Streptococcus GBS

The goal of this protocol is to describe a preclinical animal model of Group B Streptococcus (GBS)-induced chorioamnionitis. The study is designed to investigate mechanistic processes, potential causal links with developmental impairments, and finally to develop translational anti-inflammatory placento- and neuro-protective treatments.

C-section of Preclinical Animal Model of Chorioamnionitis Triggered by Group B Streptococcus (GBS)

Group B Streptococcus (GBS) is one of the most common bacteria isolated during human pregnancy. It is a leading cause of placental infection/inflammation, ...

c-section-preclinical-animal-model-chorioamnionitis-triggered-group-b

C-section of Preclinical Animal Model of Chorioamnionitis Triggered by Group B Streptococcus (GBS)

Abstract