Chlamydia muridarum infection causes bronchointerstitial pneumonia in NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice.

The murine bacterial pathogen Chlamydia muridarum (Cm) has been used to study human Chlamydia infections in various mouse models. CD4+ T-cells, natural killer cells, and interferon-gamma (IFN-γ)-mediated immunity are important to control experimentally induced Cm infections. Despite its experimental use, natural infection by Cm has not been documented in laboratory mice since the 1940s. In 2022, the authors reported the discovery of natural Cm infections in numerous academic institutional laboratory mouse colonies around the globe. To evaluate the impact of Cm infection in severely immunocompromised mice, 19 NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice were cohoused with Cm shedding, naturally infected immunocompetent mice and/or their soiled bedding for 4 weeks and subsequently euthanized. Clinical disease, characterized by lethargy, dyspnea, and weight loss, was observed in 11/19 NSG mice, and 16/18 NSG mice had neutrophilia. All mice exhibited multifocal to coalescing histiocytic and neutrophilic bronchointerstitial pneumonia (17/19) or bronchiolitis (2/19) with intraepithelial chlamydial inclusions (CIs). Immunofluorescence showed CIs were often associated with bronchiolar epithelium. CIs were frequently detected by immunohistochemistry in tracheal and bronchiolar epithelium (19/19), as well as throughout the small and large intestinal epithelium without lesions (19/19). In a subset of cases, Cm colonized the surface epithelium in the nasopharynx (16/19), nasal cavity (7/19), and middle ear canal (5/19). Endometritis and salpingitis with intraepithelial CI were identified in a single mouse. These findings demonstrate that Cm infection acquired through direct contact or soiled bedding causes significant pulmonary pathology and widespread intestinal colonization in NSG mice.

New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm(48) on the Novel Plasmid pJW2311 in Staphylococcus xylosus.

Whole-genome sequencing of Staphylococcus xylosus strain JW2311 from bovine mastitis milk identified the novel 49.3-kb macrolide-lincosamide-streptogramin B (MLSB) resistance plasmid pJW2311. It contained the macrolide resistance gene mph(C), the macrolide-streptogramin B resistance gene msr(A), and the new MLSB resistance gene erm(48) and could be transformed into Staphylococcus aureus by electroporation. Functionality of erm(48) was demonstrated by cloning and expression in S. aureus.

The new macrolide-lincosamide-streptogramin B resistance gene erm(45) is located within a genomic island in Staphylococcus fleurettii.

Genome alignment of a macrolide, lincosamide, and streptogramin B (MLSB)-resistant Staphylococcus fleurettii strain with an MLSB-susceptible S. fleurettii strain revealed a novel 11,513-bp genomic island carrying the new erythromycin resistance methylase gene erm(45). This gene was shown to confer inducible MLSB resistance when cloned into Staphylococcus aureus. The erm(45)-containing island was integrated into the housekeeping gene guaA in S. fleurettii and was able to form a circular intermediate but was not transmissible to S. aureus.

The novel macrolide-Lincosamide-Streptogramin B resistance gene erm(44) is associated with a prophage in Staphylococcus xylosus.

A novel erythromycin ribosome methylase gene, erm(44), that confers resistance to macrolide, lincosamide, and streptogramin B (MLSB) antibiotics was identified by whole-genome sequencing of the chromosome of Staphylococcus xylosus isolated from bovine mastitis milk. The erm(44) gene is preceded by a regulatory sequence that encodes two leader peptides responsible for the inducible expression of the methylase gene, as demonstrated by cloning in Staphylococcus aureus. The erm(44) gene is located on a 53-kb putative prophage designated ΦJW4341-pro. The 56 predicted open reading frames of ΦJW4341-pro are structurally organized into the five functional modules found in members of the family Siphoviridae. ΦJW4341-pro is site-specifically integrated into the S. xylosus chromosome, where it is flanked by two perfect 19-bp direct repeats, and exhibits the ability to circularize. The presence of erm(44) in three additional S. xylosus strains suggests that this putative prophage has the potential to disseminate MLSB resistance.

Evaluation of PCR electrospray-ionization mass spectrometry for rapid molecular diagnosis of bovine mastitis.

Bovine mastitis, an inflammatory disease of the mammary gland, is one of the most costly diseases affecting the dairy industry. The treatment and prevention of this disease is linked heavily to the use of antibiotics in agriculture and early detection of the primary pathogen is essential to control the disease. Milk samples (n=67) from cows suffering from mastitis were analyzed for the presence of pathogens using PCR electrospray-ionization mass spectrometry (PCR/ESI-MS) and were compared with standard culture diagnostic methods. Concurrent identification of the primary mastitis pathogens was obtained for 64% of the tested milk samples, whereas divergent results were obtained for 27% of the samples. The PCR/ESI-MS failed to identify some of the primary pathogens in 18% of the samples, but identified other pathogens as well as microorganisms in samples that were negative by culture. The PCR/ESI-MS identified bacteria to the species level as well as yeasts and molds in samples that contained a mixed bacterial culture (9%). The sensitivity of the PCR/ESI-MS for the most common pathogens ranged from 57.1 to 100% and the specificity ranged from 69.8 to 100% using culture as gold standard. The PCR/ESI-MS also revealed the presence of the methicillin-resistant gene mecA in 16.2% of the milk samples, which correlated with the simultaneous detection of staphylococci including Staphylococcus aureus. We demonstrated that PCR/ESI-MS, a more rapid diagnostic platform compared with bacterial culture, has the significant potential to serve as an important screening method in the diagnosis of bovine clinical mastitis and has the capacity to be used in infection control programs for both subclinical and clinical disease.

Outbreaks of Typhlocolitis Caused by Hypervirulent Group ST1 Clostridioides difficile in Highly Immunocompromised Strains of Mice.

Clostridioides difficile is an enteric pathogen that can cause significant clinical disease in both humans and animals. However, clinical disease arises most commonly after treatment with broad-spectrum antibiotics. The organism's ability to cause naturally occurring disease in mice is rare, and little is known about its clinical significance in highly immunocompromised mice. We report on 2 outbreaks of diarrhea associated with C. difficile in mice. In outbreak 1, 182 of approximately 2, 400 NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) and related strains of mice became clinically ill after cessation of a 14-d course of 0.12% amoxicillin feed to control an increase in clinical signs associated with Corynebacterium bovis infection. Most mice had been engrafted with human tumors; the remainder were experimentally naïve. Affected animals exhibited 1 of 3 clinical syndromes: 1) peracute death; 2) severe diarrhea leading to euthanasia or death; or 3) mild to moderate diarrhea followed by recovery. A given cage could contain both affected and unaffected mice. Outbreak 2 involved a small breeding colony (approximately 50 mice) of NOD. CB17-Prkdcscid/NCrCrl (NOD-scid) mice that had not received antibiotics or experimental manipulations. In both outbreaks, C. difficile was isolated, and toxins A and B were detected in intestinal content or feces. Histopathologic lesions highly suggestive of C. difficile enterotoxemia included fibrinonecrotizing and neutrophilic typhlocolitis with characteristic 'volcano' erosions or pseudomembrane formation. Genomic analysis of 4 isolates (3 from outbreak 1 and 1 from outbreak 2) revealed that these isolates were closely related to a pathogenic human isolate, CD 196. To our knowledge, this report is the first to describe naturally occurring outbreaks of C. difficile-associated typhlocolitis with significant morbidity and mortality in highly immunocompromised strains of mice.

Genome Sequences of Six Prophages Isolated from Staphylococcus pseudintermedius Strains Recovered from Human and Animal Clinical Specimens.

Staphylococcus pseudintermedius is a common bacterial pathogen in companion animal medicine and has demonstrated zoonotic potential. Here, we report six new Staphylococcus pseudintermedius prophage genomes of the Siphoviridae family, identified in isolates recovered from human and canine clinical specimens.