IKK2/NFkB signaling controls lung resident CD8+ T cell memory during influenza infection.

CD8+ T cell tissue resident memory (TRM) cells are especially suited to control pathogen spread at mucosal sites. However, their maintenance in lung is short-lived. TCR-dependent NFkB signaling is crucial for T cell memory but how and when NFkB signaling modulates tissue resident and circulating T cell memory during the immune response is unknown. Here, we find that enhancing NFkB signaling in T cells once memory to influenza is established, increases pro-survival Bcl-2 and CD122 levels thus boosting lung CD8+ TRM maintenance. By contrast, enhancing NFkB signals during the contraction phase of the response leads to a defect in CD8+ TRM differentiation without impairing recirculating memory subsets. Specifically, inducible activation of NFkB via constitutive active IKK2 or TNF interferes with TGFß signaling, resulting in defects of lung CD8+ TRM imprinting molecules CD69, CD103, Runx3 and Eomes. Conversely, inhibiting NFkB signals not only recovers but improves the transcriptional signature and generation of lung CD8+ TRM. Thus, NFkB signaling is a critical regulator of tissue resident memory, whose levels can be tuned at specific times during infection to boost lung CD8+ TRM.

Disruption of the Francisella noatunensis subsp. orientalis pdpA Gene Results in Virulence Attenuation and Protection in Zebrafish.

Several Francisella spp., including Francisella noatunensis, are regarded as important emerging pathogens of wild and farmed fish. However, very few studies have investigated the virulence factors that allow these bacterial species to be pathogenic in fish. The Francisella pathogenicity island (FPI) is a well-described, gene-dense region encoding major virulence factors for the genus Francisella. pdpA is a member of the pathogenicity-determining protein genes carried by the FPI that are implicated in the ability of the mammalian pathogen Francisella tularensis to escape and replicate in infected host cells. Using a sacB suicide approach, we generated pdpA knockouts to address the role of PdpA as a virulence factor for F. noatunensis. Because polarity can be an issue in gene-dense regions, we generated two different marker-based mutants in opposing polarity (the F. noatunensis subsp. orientalis ΔpdpA1 and ΔpdpA2 strains). Both mutants were attenuated (P < 0.0001) in zebrafish challenges and displayed impaired intracellular replication (P < 0.05) and cytotoxicity (P < 0.05), all of which could be restored to wild-type (WT) levels by complementation for the ΔpdpA1 mutant. Importantly, differences were found for bacterial burden and induction of acute-phase and proinflammatory genes for the F. noatunensis subsp. orientalis ΔpdpA1 and ΔpdpA2 mutants compared to the WT during acute infection. In addition, neither mutant resulted in significant histopathological changes. Finally, immunization with the F. noatunensis subsp. orientalis ΔpdpA1 mutant led to protection (P < 0.012) against an acute 40% lethal dose (LD40) challenge with WT F. noatunensis in the zebrafish model of infection. Taken together, the results from this study further demonstrate physiological similarities within the genus Francisella relative to their phylogenetic relationships and the utility of zebrafish for addressing virulence factors for the genus.

Whole-Genome Comparisons of Staphylococcus agnetis Isolates from Cattle and Chickens.

Staphylococcus agnetis has been previously associated with subclinical or clinically mild cases of mastitis in dairy cattle and is one of several staphylococcal species that have been isolated from the bones and blood of lame broilers. We reported that S. agnetis could be obtained frequently from bacterial chondronecrosis with osteomyelitis (BCO) lesions of lame broilers (A. Al-Rubaye et al., PLoS One 10:e0143336, 2015 [https://doi.org/10.1371/journal.pone.0143336]). A particular isolate, S. agnetis 908, can induce lameness in over 50% of exposed chickens, exceeding normal BCO incidences in broiler operations. We reported the assembly and annotation of the genome of isolate 908. To better understand the relationship between dairy cattle and broiler isolates, we assembled 11 additional genomes for S. agnetis isolates, an additional chicken BCO strain, and ten isolates from cattle milk, mammary gland secretions, or udder skin from the collection at the University of Missouri. To trace phylogenetic relationships, we constructed phylogenetic trees based on multilocus sequence typing and genome-to-genome distance comparisons. Chicken isolate 908 clustered with two of the cattle isolates, along with three isolates from chickens in Denmark and an isolate of S. agnetis we isolated from a BCO lesion on a commercial broiler farm in Arkansas. We used a number of BLAST tools to compare the chicken isolates to those from cattle and identified 98 coding sequences distinguishing isolate 908 from the cattle isolates. None of the identified genes explain the differences in host or tissue tropism. These analyses are critical to understanding how staphylococci colonize and infect different hosts and potentially how they can transition to alternative niches (bone versus dermis).IMPORTANCEStaphylococcus agnetis has been recently recognized as associated with disease in dairy cattle and meat-type chickens. The infections appear to be limited in cattle and systemic in broilers. This report details the molecular relationships between cattle and chicken isolates in order to understand how this recently recognized species infects different hosts with different disease manifestations. The data show that the chicken and cattle isolates are very closely related, but the chicken isolates all cluster together, suggesting a single jump from cattle to chickens.

An outbreak of Mycoplasma mycoides subspecies capri arthritis in young goats: a case study.

Mycoplasmosis is a well-known cause of morbidity and mortality in small ruminants. Previously recognized outbreaks have involved arthritis, and pneumonia or pleuropneumonia. Modern bacteriology procedures rely less on isolation techniques that require special media for mollicutes given that these species are notoriously difficult to isolate, and rely more on PCR tests. We report an outbreak of arthritis, pleuropneumonia, and mild meningitis affecting dairy goat kids, spanning a period of 3 y, which had unusual epidemiologic characteristics related to husbandry practices. Lesions were characterized by polyarthritis of the appendicular joints, with copious joint fluid and extension of arthritic exudate beyond the joint itself. The cause remained unknown until serendipitous isolation of a mycoplasma on blood agar. Mycoplasmosis was not detected from synovial samples by a general mycoplasma PCR, despite multiple attempts. Isolated colonies were also negative by this general PCR assay. The isolate was identified as Mycoplasma mycoides subspecies capri, using universal 16S primers and amplicon sequencing. Testing of additional isolates from other diseased goats in the herd confirmed that this was the cause of illness. A failure to recognize the distinct nature of organisms of the M. mycoides group of mycoplasmas meant that a PCR test that cannot detect this group of organisms was utilized at first, and the etiology of the illness was overlooked for a period of time. Veterinary pathologists and microbiologists must be aware of the limitations of some PCR assays when confronted with joint disease and pleuropneumonia in small ruminants.

Staphylococcal intramammary infection dynamics and the relationship with milk quality parameters in dairy goats over the dry period.

The objectives of this study were (1) to report the rates of new intramammary infection (IMI) and spontaneous IMI cure over the dry period in 3 dairy goat herds; (2) to evaluate the factors predicting infection dynamics over the dry period; and (3) to define milk quality parameter thresholds that predict infection dynamics over the dry period. Two consecutive udder-half milk samples were collected 10 to 14 d apart before dry-off from 288 goats in 3 herds, and 2 consecutive udder-half samples were collected 7 to 14 d apart in the following lactation, with the first sample being collected ≤10 d in milk, from 200 of the same goats. In 2 of the herds, udder-half milk samples were also collected at the same time points (n = 312 halves; 157 goats) for measurement of milk quality parameters. Standard aerobic culture of milk samples was performed for the detection of mastitis pathogens. To rule out the presence of Mycoplasma spp. IMI, milk samples were also cultured on modified Hayflick medium. Non-Mycoplasma isolates were speciated using MALDI-TOF mass spectrometry. Staphylococcal isolates, when not identified by MALDI-TOF, were speciated using partial gene sequence analysis of rpoB or tuf. When >1 sample from an udder half yielded the same species, available isolates from the first and last positive samples for that species were strain-typed using pulsed-field gel electrophoresis. Incidence of new IMI and cure rate were computed. Generalized linear mixed regression models were built to evaluate the associations between new IMI and pre-dry somatic cell score (SCS), between IMI persistence and half-level SCS, and between IMI persistence and pre-dry IMI species. Thresholds for pre-dry SCS and lactose concentration were computed to predict IMI persistence. Overall, 12.6% (48/380) of halves had a persistent IMI. Cumulative incidence of new IMI over the dry period was 13.2%, and cure rate was 52.0%. Pre-dry SCS was not associated with odds of new IMI or IMI persistence. Pre-dry IMI species was not associated with odds of persistence. Lactose concentration was not associated with odds of persistence. Regardless of culture data, the optimal pre-dry SCS threshold to detect IMI that would persist into the next lactation was 8.7, with sensitivity and specificity of 50 and 73.8%, respectively. Further studies on the effect of control measures on species-specific incidence and cure rates during the dry period are warranted.

INTESTINAL HISTOPLASMOSIS IN A CAPTIVE REINDEER (RANGIFER TARANDUS), MISSOURI, USA.

An infection with Histoplasma capsulatum was diagnosed in a farmed reindeer in Missouri, an endemic area for histoplasmosis, localized in the intestine. The intrahistiocytic organisms were identified in tissue sections using histologic methods and confirmed by immunohistochemistry. This is the first report of histoplasmosis in a reindeer or in any deer species.

Complete Genome Sequence of Kocuria palustris MU14/1.

Presented here is the first completely assembled genome sequence of Kocuria palustris, an actinobacterial species with broad ecological distribution. The single, circular chromosome of K. palustris MU14/1 comprises 2,854,447 bp, has a G+C content of 70.5%, and contains a deduced gene set of 2,521 coding sequences.

Genome Sequence Analysis of Mycoplasma sp. HU2014, Isolated from Tissue Culture.

The draft genome sequence of a novel Mycoplasma strain, designated Mycoplasma sp. HU2014, has been determined. The genome comprises 1,084,927 nucleotides and was obtained from a mycoplasma-infected culture of chicken DT40 cells. Phylogenetic analysis places this taxon in a group comprising the closely related species Mycoplasma yeatsii and Mycoplasma cottewii.

Complete Genome Sequence of Mycoplasma hominis Strain Sprott (ATCC 33131), Isolated from a Patient with Nongonococcal Urethritis.

Presented here is the complete and annotated genome sequence of Mycoplasma hominis Sprott (ATCC 33131). The chromosome comprises 695,214 bp, which is approximately 30 kb larger than the syntenic genome of M. hominis PG21(T). Tetracycline resistance of strain Sprott is most probably conferred by the tetM determinant, harbored on a mosaic transposon-like structure.

Comparative analysis of the Mycoplasma capricolum subsp. capricolum GM508D genome reveals subrogation of phase-variable contingency genes and a novel integrated genetic element.

Mycoplasma capricolum subspecies capricolum is both a pathogen of small ruminants and a model recipient organism for gene transplantation and synthetic biology. With the availability of the complete genome of the type strain California kid (released in 2005), a draft genome of strain GM508D was determined to investigate genomic variation in this subspecies. Differences in mobile genetic element location and complement, catabolic pathway genes, contingency loci, surface antigen genes and type II restriction-modification systems highlight the plasticity and diversity within this taxon.

Complete Genome Sequence of Mycoplasma yeatsii Strain GM274B (ATCC 43094).

Mycoplasma yeatsii is a goat mycoplasma species that, although an obligate parasite, accommodates this lifestyle as an inapparent commensalist. High-frequency transformation has also been reported for this species. The complete 895,051-bp genome sequence of strain GM274B has been determined, enabling an analysis of the features of this potential cloning host.

Complete Genome Sequence of Mycoplasma flocculare Strain Ms42T (ATCC 27399T).

Mycoplasma flocculare is a commensal or low-virulence pathogen of swine. The complete 778,866-bp genome sequence of M. flocculare strain Ms42(T) has been determined, enabling further comparison to genomes of the closely related pathogen Mycoplasma hyopneumoniae. The absence of the p97 and glpD genes may contribute to the attenuated virulence of M. flocculare.

Sequence Analysis of Staphylococcus hyicus ATCC 11249T, an Etiological Agent of Exudative Epidermitis in Swine, Reveals a Type VII Secretion System Locus and a Novel 116-Kilobase Genomic Island Harboring Toxin-Encoding Genes.

Staphylococcus hyicus is the primary etiological agent of exudative epidermitis in swine. Analysis of the complete genome sequence of the type strain revealed a locus encoding a type VII secretion system and a large chromosomal island harboring the genes encoding exfoliative toxin ExhA and an EDIN toxin homolog.

Analysis of the Complete Mycoplasma hominis LBD-4 Genome Sequence Reveals Strain-Variable Prophage Insertion and Distinctive Repeat-Containing Surface Protein Arrangements.

The complete genome sequence of Mycoplasma hominis LBD-4 has been determined and the gene content ascribed. The 715,165-bp chromosome contains 620 genes, including 14 carried by a strain-variable prophage genome related to Mycoplasma fermentans MFV-1 and Mycoplasma arthritidis MAV-1. Comparative analysis with the genome of M. hominis PG21(T) reveals distinctive arrangements of repeat-containing surface proteins.

Isolation of Aureimonas altamirensis, a Brucella canis-like bacterium, from an edematous canine testicle.

Microbiological and histological analysis of a sample from a swollen testicle of a 2-year-old Border Collie dog revealed a mixed infection of the fungus Blastomyces dermatitidis and the Gram-negative bacterium Aureimonas altamirensis. When subjected to an automated microbial identification system, the latter isolate was provisionally identified as Psychrobacter phenylpyruvicus, but the organism shared several biochemical features with Brucella canis and exhibited agglutination, albeit weakly, with anti-B. canis antiserum. Unequivocal identification of the organism was only achieved by 16S ribosomal RNA gene sequencing, ultimately establishing the identity as A. altamirensis. Since its first description in 2006, this organism has been isolated infrequently from human clinical samples, but, to the authors' knowledge, has not been reported from a veterinary clinical sample. While of unknown clinical significance with respect to the pathology observed for the polymicrobial infection described herein, it highlights the critical importance to unambiguously identify the microbe for diagnostic, epidemiological, infection control, and public health purposes.

Genome Sequence of Actinobacillus suis Type Strain ATCC 33415T.

The assembled and annotated genome of Actinobacillus suis ATCC 33415(T) is reported here. The 2,501,598-bp genome encodes 2,246 open reading frames (ORFs) with strain variable incursion of an integrative conjugative element into a tRNA locus. Comparative analysis of the deduced gene set should inform our understanding of pathogenesis, genomic plasticity, and serotype variation.

Draft Genome Sequence of Bovine Mastitis Isolate Staphylococcus agnetis CBMRN 20813338.

Presented here is a draft genome sequence for Staphylococcus agnetis CBMRN 20813338, isolated from a lactating dairy cow with subclinical mastitis. The genome is approximately 2,416 kb and has 35.79% G+C content. Analysis of the deduced open reading frame (ORF) set identified candidate virulence attributes in addition to potential molecular targets for species identification.

Draft Genome Sequence of Staphylococcus chromogenes Strain MU 970, Isolated from a Case of Chronic Bovine Mastitis.

Coagulase-negative staphylococcal species are a common cause of subclinical bovine mastitis, with Staphylococcus chromogenes being one of the most frequently identified species in these cases. The draft genome sequence of an S. chromogenes isolate (MU 970) recovered from the milk of a cow with a chronic intramammary infection is reported here.