Anti-biofilm Activity of Human Milk Oligosaccharides in Clinical Strains of Streptococcus agalactiae with Diverse Capsular and Sequence Types.

Group B Streptococcus (GBS) is an encapsulated Gram-positive bacterial pathogen that causes severe perinatal infections. Human milk oligosaccharides (HMOs) are short-chain sugars that have recently been shown to possess antimicrobial and anti-biofilm activity against a variety of bacterial pathogens, including GBS. We have expanded these studies to demonstrate that HMOs can inhibit and dismantle biofilm in both invasive and colonizing strains of GBS. A cohort of 30 diverse strains of GBS were analyzed for susceptibility to HMO-dependent biofilm inhibition or destruction. HMOs were significantly effective at inhibiting biofilm in capsular-type- and sequence-type-specific fashion, with significant efficacy in CpsIb, CpsII, CpsIII, CpsV, and CpsVI strains as well as ST-1, ST-12, ST-19, and ST-23 strains. Interestingly, CpsIa as well as ST-7 and ST-17 were not susceptible to the anti-biofilm activity of HMOs, underscoring the strain-specific effects of these important antimicrobial molecules against the perinatal pathogen Streptococcus agalactiae.

Galacto-Oligosaccharide Supplementation Modulates Pathogen-Commensal Competition between Streptococcus agalactiae and Streptococcus salivarius.

The members of the infant microbiome are governed by feeding method (breastmilk vs. formula). Regardless of the source of nutrition, a competitive growth advantage can be provided to commensals through prebiotics - either human milk oligosaccharides (HMOs) or plant oligosaccharides that are supplemented into formula. To characterize how prebiotics modulate commensal - pathogen interactions, we have designed and studied a minimal microbiome where a pathogen, Streptococcus agalactiae engages with a commensal, Streptococcus salivarius. We discovered that while S. agalactiae suppresses the growth of S. salivarius via increased lactic acid production, galacto-oligosaccharides (GOS) supplementation reverses the effect. This result has major implications in characterizing how single species survive in the gut, what niche they occupy, and how they engage with other community members.

The antimicrobial activity of zinc against group B Streptococcus is strain-dependent across diverse sequence types, capsular serotypes, and invasive versus colonizing isolates.

BACKGROUND: Streptococcus agalactiae or Group B Streptococcus (GBS) is an encapsulated gram-positive bacterial pathobiont that commonly colonizes the lower gastrointestinal tract and reproductive tract of human hosts. This bacterium can infect the gravid reproductive tract and cause invasive infections of pregnant patients and neonates. Upon colonizing the reproductive tract, the bacterial cell is presented with numerous nutritional challenges imposed by the host. One strategy employed by the host innate immune system is intoxication of bacterial invaders with certain transition metals such as zinc. METHODOLOGY: Previous work has demonstrated that GBS must employ elegant strategies to circumnavigate zinc stress in order to survive in the vertebrate host. We assessed 30 strains of GBS from diverse isolation sources, capsular serotypes, and sequence types for susceptibility or resistance to zinc intoxication. RESULTS: Invasive strains, such as those isolated from early onset disease manifestations of GBS infection were significantly less susceptible to zinc toxicity than colonizing strains isolated from rectovaginal swabs of pregnant patients. Additionally, capsular type III (cpsIII) strains and the ST-17 and ST-19 strains exhibited the greatest resilience to zinc stress, whereas ST-1 and ST-12 strains as well as those possessing capsular type Ib (cpsIb) were more sensitive to zinc intoxication. Thus, this study demonstrates that the transition metal zinc possesses antimicrobial properties against a wide range of GBS strains, with isolation source, capsular serotype, and sequence type contributing to susceptibility or resistance to zinc stress.

Distinct Group B Streptococcus Sequence and Capsule Types Differentially Impact Macrophage Stress and Inflammatory Signaling Responses.

Group B Streptococcus (GBS) is an opportunistic bacterial pathogen that can contribute to the induction of preterm birth in colonized pregnant women and to severe neonatal disease. Many questions regarding the mechanisms that drive GBS-associated pathogenesis remain unanswered, and it is not yet clear why virulence has been observed to vary so extensively across GBS strains. Previously, we demonstrated that GBS strains of different sequence types (STs) and capsule (CPS) types induce different cytokine profiles in infected THP-1 macrophage-like cells. Here, we expanded on these studies by utilizing the same set of genetically diverse GBS isolates to assess ST and CPS-specific differences in upstream cell death and inflammatory signaling pathways. Our results demonstrate that particularly virulent STs and CPS types, such as the ST-17 and CPS III groups, induce enhanced Jun-N-terminal protein kinase (JNK) and NF-κB pathway activation following GBS infection of macrophages compared with other ST or CPS groups. Additionally, we found that ST-17, CPS III, and CPS V GBS strains induce the greatest levels of macrophage cell death during infection and exhibit a more pronounced ability to be internalized and to survive in macrophages following phagocytosis. These data provide further support for the hypothesis that variable host innate immune responses to GBS, which significantly impact pathogenesis, stem in part from genotypic and phenotypic differences among GBS isolates. These and similar studies may inform the development of improved diagnostic, preventive, or therapeutic strategies targeting invasive GBS infections.

Antibiotic Susceptibility Profiles and Frequency of Resistance Genes in Clinical Shiga Toxin-Producing Escherichia coli Isolates from Michigan over a 14-Year Period.

Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen that contributes to over 250,000 infections in the United States each year. Because antibiotics are not recommended for STEC infections, resistance in STEC has not been widely researched despite an increased likelihood for the transfer of resistance genes from STEC to opportunistic pathogens residing within the same microbial community. From 2001 to 2014, 969 STEC isolates were collected from Michigan patients. Antibiotic susceptibility profiles to clinically relevant antibiotics were determined using disc diffusion, while epidemiological data were used to identify factors associated with resistance. Whole-genome sequencing was used for serotyping, examining genetic relatedness, and identifying genetic determinants and mechanisms of resistance in the non-O157 isolates. Increasing frequencies of resistance to at least one antibiotic were observed over the 14 years (P = 0.01). While the non-O157 serogroups were more commonly resistant than O157 (odds ratio, 2.4; 95% confidence interval,1.43 to 4.05), the frequency of ampicillin resistance among O157 isolates was significantly higher in Michigan than the national average (P = 0.03). Genomic analysis of 321 non-O157 isolates uncovered 32 distinct antibiotic resistance genes (ARGs). Although mutations in genes encoding resistance to ciprofloxacin and ampicillin were detected in four isolates, most of the horizontally acquired ARGs conferred resistance to aminoglycosides, ß-lactams, sulfonamides, and/or tetracycline. This study provides insight into the mechanisms of resistance in a large collection of clinical non-O157 STEC isolates and demonstrates that antibiotic resistance among all STEC serogroups has increased over time, prompting the need for enhanced surveillance.

Antibacterial and Anti-biofilm Activity of the Human Breast Milk Glycoprotein Lactoferrin against Group B Streptococcus.

Group B Streptococcus (GBS) is an encapsulated Gram-positive human pathogen that causes invasive infections in pregnant hosts and neonates, as well as immunocompromised individuals. Colonization of the human host requires the ability to adhere to mucosal surfaces and circumnavigate the nutritional challenges and antimicrobial defenses associated with the innate immune response. Biofilm formation is a critical process to facilitate GBS survival and establishment of a replicative niche in the vertebrate host. Previous work has shown that the host responds to GBS infection by producing the innate antimicrobial glycoprotein lactoferrin, which has been implicated in repressing bacterial growth and biofilm formation. Additionally, lactoferrin is highly abundant in human breast milk and could serve a protective role against invasive microbial pathogens. This study demonstrates that human breast milk lactoferrin has antimicrobial and anti-biofilm activity against GBS and inhibits its adherence to human gestational membranes. Together, these results indicate that human milk lactoferrin could be used as a prebiotic chemotherapeutic strategy to limit the impact of bacterial adherence and biofilm formation on GBS-associated disease outcomes.

Analysis of virulence phenotypes and antibiotic resistance in clinical strains of Acinetobacter baumannii isolated in Nashville, Tennessee.

BACKGROUND: Acinetobacter baumannii is a gram-negative bacterium which causes opportunistic infections in immunocompromised hosts. Genome plasticity has given rise to a wide range of strain variation with respect to antimicrobial resistance profiles and expression of virulence factors which lead to altered phenotypes associated with pathogenesis. The purpose of this study was to analyze clinical strains of A. baumannii for phenotypic variation that might correlate with virulence phenotypes, antimicrobial resistance patterns, or strain isolation source. We hypothesized that individual strain virulence phenotypes might be associated with anatomical site of isolation or alterations in susceptibility to antimicrobial interventions. METHODOLOGY: A cohort of 17 clinical isolates of A. baumannii isolated from diverse anatomical sites were evaluated to ascertain phenotypic patterns including biofilm formation, hemolysis, motility, and antimicrobial resistance. Antibiotic susceptibility/resistance to ampicillin-sulbactam, amikacin, ceftriaxone, ceftazidime, cefotaxime, ciprofloxacin, cefepime, gentamicin, levofloxacin, meropenem, piperacillin, trimethoprim-sulfamethoxazole, ticarcillin- K clavulanate, tetracyclin, and tobramycin was determined. RESULTS: Antibiotic resistance was prevalent in many strains including resistance to ampicillin-sulbactam, amikacin, ceftriaxone, ceftazidime, cefotaxime, ciprofloxacin, cefepime, gentamicin, levofloxacin, meropenem, piperacillin, trimethoprim-sulfamethoxazole, ticarcillin- K clavulanate, tetracyclin, and tobramycin. All strains tested induced hemolysis on agar plate detection assays. Wound-isolated strains of A. baumannii exhibited higher motility than strains isolated from blood, urine or Foley catheter, or sputum/bronchial wash. A. baumannii strains isolated from patient blood samples formed significantly more biofilm than isolates from wounds, sputum or bronchial wash samples. An inverse relationship between motility and biofilm formation was observed in the cohort of 17 clinical isolates of A. baumannii tested in this study. Motility was also inversely correlated with induction of hemolysis. An inverse correlation was observed between hemolysis and resistance to ticarcillin-k clavulanate, meropenem, and piperacillin. An inverse correlation was also observed between motility and resistance to ampicillin-sulbactam, ceftriaxone, ceftoxamine, ceftazidime, ciprofloxacin, or levofloxacin. CONCLUSIONS: Strain dependent variations in biofilm and motility are associated with anatomical site of isolation. Biofilm and hemolysis production both have an inverse association with motility in the cohort of strains utilized in this study, and motility and hemolysis were inversely correlated with resistance to numerous antibiotics.

Population Gene Introgression and High Genome Plasticity for the Zoonotic Pathogen Streptococcus agalactiae.

The influence that bacterial adaptation (or niche partitioning) within species has on gene spillover and transmission among bacterial populations occupying different niches is not well understood. Streptococcus agalactiae is an important bacterial pathogen that has a taxonomically diverse host range making it an excellent model system to study these processes. Here, we analyze a global set of 901 genome sequences from nine diverse host species to advance our understanding of these processes. Bayesian clustering analysis delineated 12 major populations that closely aligned with niches. Comparative genomics revealed extensive gene gain/loss among populations and a large pan genome of 9,527 genes, which remained open and was strongly partitioned among niches. As a result, the biochemical characteristics of 11 populations were highly distinctive (significantly enriched). Positive selection was detected and biochemical characteristics of the dispensable genes under selection were enriched in ten populations. Despite the strong gene partitioning, phylogenomics detected gene spillover. In particular, tetracycline resistance (which likely evolved in the human-associated population) from humans to bovine, canines, seals, and fish, demonstrating how a gene selected in one host can ultimately be transmitted into another, and biased transmission from humans to bovines was confirmed with a Bayesian migration analysis. Our findings show high bacterial genome plasticity acting in balance with selection pressure from distinct functional requirements of niches that is associated with an extensive and highly partitioned dispensable genome, likely facilitating continued and expansive adaptation.

Genetic and Phenotypic Factors Associated with Persistent Shedding of Shiga Toxin-Producing Escherichia coli by Beef Cattle.

Shiga toxin-producing Escherichia coli (STEC) is a leading cause of foodborne infections. Cattle are an important STEC reservoir, although little is known about specific pathogen traits that impact persistence in the farm environment. Hence, we sought to evaluate STEC isolates recovered from beef cattle in a single herd in Michigan. To do this, we collected fecal grabs from 26 cattle and resampled 13 of these animals at 3 additional visits over a 3-month period. In all, 66 STEC isolates were recovered for genomics and biofilm quantification using crystal violet assays. The STEC population was diverse, representing seven serotypes, including O157:H7, O26:H11, and O103:H2, which are commonly associated with human infections. Although a core genome analysis of 2,933 genes grouped isolates into clusters based on serogroups, some isolates within each cluster had variable biofilm levels and virulence gene profiles. Most (77.8%; n = 49) isolates harbored stx2a, while 38 (57.5%) isolates formed strong biofilms. Isolates belonging to the predominant serogroup O6 (n = 36; 54.5%) were more likely to form strong biofilms, persistently colonize multiple cattle, and be acquired over time. A high-quality single nucleotide polymorphism (SNP) analysis of 33 O6 isolates detected between 0 and 13 single nucleotide polymorphism (SNP) differences between strains, indicating that highly similar strain types were persisting in this herd. Similar findings were observed for other persistent serogroups, although key genes were found to differ among strong and weak biofilm producers. Together, these data highlight the diversity and persistent nature of some STEC types in this important food animal reservoir.IMPORTANCE Food animal reservoirs contribute to Shiga toxin-producing Escherichia coli (STEC) evolution via the acquisition of horizontally acquired elements like Shiga toxin bacteriophages that enhance pathogenicity. In cattle, persistent fecal shedding of STEC contributes to contamination of beef and dairy products and to crops being exposed to contaminated water systems. Hence, identifying factors important for STEC persistence is critical. This longitudinal study enhances our understanding of the genetic diversity of STEC types circulating in a cattle herd and identifies genotypic and phenotypic traits associated with persistence. Key findings demonstrate that multiple STEC types readily persist in and are transmitted across cattle in a shared environment. These dynamics also enhance the persistence of virulence genes that can be transferred between bacterial hosts, resulting in the emergence of novel STEC strain types. Understanding how pathogens persist and diversify in reservoirs is important for guiding new preharvest prevention strategies aimed at reducing foodborne transmission to humans.

The impact of Lactobacillus on group B streptococcal interactions with cells of the extraplacental membranes.

Group B Streptococcus (GBS) causes adverse pregnancy outcomes and neonatal disease. The recommended preventative measure is intrapartum antibiotic prophylaxis, which can prevent early onset neonatal disease but not chorioamnionitis, preterm labor, stillbirth, or late-onset disease. Novel prevention methods are therefore needed. Use of probiotics including Lactobacillus spp., has been suggested given that they are dominant members of the lower reproductive tract microbiome. Although Lactobacillus was shown to reduce recto-vaginal colonization of GBS, no studies have examined how Lactobacillus impacts GBS in the extraplacental membranes. Since Lactobacillus has been detected in the placental membranes, we sought to characterize GBS-Lactobacillus interactions in vitro using a colonizing and invasive GBS strain. While live Lactobacillus did not affect growth or biofilms in GBS, co-culture with L. gasseri led to a 224-fold increase in GBS association with decidualized human endometrial stromal cells for both GBS strains (p < 0.005). Increased association did not result in increased invasion (p > 0.05) or host cell death, though some GBS and Lactobacillus combinations contributed to a significant reduction in host cell death (p < 0.05). Since Lactobacillus secretes many inhibitory compounds, the effect of Lactobacillus supernatants on GBS was also examined. The supernatants inhibited GBS growth, biofilm formation and invasion of host cells, though strain dependent effects were observed. Notably, supernatant from L. reuteri 6475 broadly inhibited growth in 36 distinct GBS strains and inhibited GBS growth to an average of 46.6% of each GBS strain alone. Together, these data show that specific Lactobacillus strains and their secreted products have varying effects on GBS interactions with cells of the extraplacental membranes that could impact pathogenesis. Understanding these interactions could help guide new treatment options aimed at reducing GBS-associated maternal complications and disease.

Modulation of Death and Inflammatory Signaling in Decidual Stromal Cells following Exposure to Group B Streptococcus.

Group B Streptococcus (GBS) is an opportunistic bacterial pathogen that contributes to miscarriage, preterm birth, and serious neonatal infections. Studies have indicated that some multilocus sequence types (STs) of GBS are more likely to cause severe disease than others. We hypothesized that the ability of GBS to elicit varying host responses in maternal decidual tissue during pregnancy is an important factor regulating infection and disease severity. To address this hypothesis, we utilized an antibody microarray to compare changes in production and activation of host signaling proteins in decidualized telomerase-immortalized human endometrial stromal cells (dT-HESCs) following infection with GBS strains from septic neonates or colonized mothers. GBS infection increased levels of total and phosphorylated mitogen-activated protein kinase (MAPK) family members such as p38 and JNK and induced nuclear factor kappa B (NF-κB) pathway activation. Infection also altered the regulation of additional proteins that mediate cell death and inflammation in a strain-specific manner, which could be due to the observed variation in attachment to and invasion of the decidual stromal cells and ability to lyse red blood cells. Further analyses confirmed array results and revealed that p38 promotes programmed necrosis in dT-HESCs. Together, the observed signaling changes may contribute to deregulation of critical developmental signaling cascades and inflammatory responses following infection, both of which could trigger GBS-associated pregnancy complications.

Intrinsic Maturational Neonatal Immune Deficiencies and Susceptibility to Group B Streptococcus Infection.

Although a normal member of the gastrointestinal and vaginal microbiota, group B Streptococcus (GBS) can also occasionally be the cause of highly invasive neonatal disease and is an emerging pathogen in both elderly and immunocompromised adults. Neonatal GBS infections are typically transmitted from mother to baby either in utero or during passage through the birth canal and can lead to pneumonia, sepsis, and meningitis within the first few months of life. Compared to the adult immune system, the neonatal immune system has a number of deficiencies, making neonates more susceptible to infection. Recognition of GBS by the host immune system triggers an inflammatory response to clear the pathogen. However, GBS has developed several mechanisms to evade the host immune response. A comprehensive understanding of this interplay between GBS and the host immune system will aid in the development of new preventative measures and therapeutics.

A Nonhemolytic Group B Streptococcus Strain Exhibits Hypervirulence.

Group B streptococci (GBS) are Gram-positive bacteria that are a leading cause of neonatal infections. Most invasive isolates are ß-hemolytic, and hemolytic activity is critical for GBS virulence. Although nonhemolytic GBS strains are occasionally isolated, they are often thought to be virulence attenuated. In this study, we show that a nonhemolytic GBS strain (GB37) isolated from a septic neonate exhibits hypervirulence. Substitution of tryptophan to leucine (W297L) in the sensor histidine kinase CovS results in constitutive kinase signaling, leading to decreased hemolysis and increased activity of the GBS hyaluronidase, HylB. These results describe how nonpigmented and nonhemolytic GBS strains can exhibit hypervirulence.

Antimicrobial Drug-Resistant Shiga Toxin-Producing Escherichia coli Infections, Michigan, USA.

High frequencies of antimicrobial drug resistance were observed in O157 and non-O157 Shiga toxin-producing E. coli strains recovered from patients in Michigan during 2010-2014. Resistance was more common in non-O157 strains and independently associated with hospitalization, indicating that resistance could contribute to more severe disease outcomes.

Intestinal Microbial Community Dynamics of White-Tailed Deer (Odocoileus virginianus) in an Agroecosystem.

The intestinal microbiota has important functions that contribute to host health. The compositional dynamics of microbial communities are affected by many factors, including diet and presence of pathogens. In contrast to humans and domestic mammals, the composition and seasonal dynamics of intestinal microbiota of wildlife species remain comparatively understudied. White-tailed deer (Odocoileus virginianus) is an ecologically and economically important wildlife species that inhabits agricultural ecosystems and is known to be a reservoir of enteric pathogens. Nevertheless, there is a lack of knowledge of white-tailed deer intestinal microbiota diversity and taxonomic composition. This study's first objective was to characterize and compare the intestinal microbiota of 66 fecal samples from white-tailed deer collected during two sampling periods (March and June) using 16S rDNA pyrosequencing. Associations between community diversity and composition and factors including season, sex, host genetic relatedness, and spatial location were quantified. Results revealed that white-tailed deer intestinal microbiota was predominantly comprised of phyla Firmicutes and Proteobacteria, whose relative frequencies varied significantly between sampling periods. The second objective was to examine the associations between the presence of Escherichia coli and Salmonella, and microbiota composition and diversity. Results indicated that relative abundance of some microbial taxa varied when a pathogen was present. This study provides insights into microbial compositional dynamics of a wildlife species inhabiting coupled natural and agricultural landscapes. Data focus attention on the high prevalence of Proteobacteria particularly during the summer and highlight the need for future research regarding the role of white-tailed deer as a natural pathogen reservoir in agroecosystems.

Factors Associated with Shiga Toxin-Producing Escherichia coli Shedding by Dairy and Beef Cattle.

UNLABELLED: Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen that can cause hemorrhagic colitis and hemolytic-uremic syndrome. Cattle are the primary reservoir for STEC, and food or water contaminated with cattle feces is the most common source of infections in humans. Consequently, we conducted a cross-sectional study of 1,096 cattle in six dairy herds (n = 718 animals) and five beef herds (n = 378 animals) in the summers of 2011 and 2012 to identify epidemiological factors associated with shedding. Fecal samples were obtained from each animal and cultured for STEC. Multivariate analyses were performed to identify risk factors associated with STEC positivity. The prevalence of STEC was higher in beef cattle (21%) than dairy cattle (13%) (odds ratio [OR], 1.76; 95% confidence interval [CI], 1.25, 2.47), with considerable variation occurring across herds (range, 6% to 54%). Dairy cattle were significantly more likely to shed STEC when the average temperature was >28.9°C 1 to 5 days prior to sampling (OR, 2.5; 95% CI, 1.25, 4.91), during their first lactation (OR, 1.8; 95% CI, 1.1, 2.8), and when they were <30 days in milk (OR, 3.9; 95% CI, 2.1, 7.2). These data suggest that the stress or the negative energy balance associated with lactation may result in increased STEC shedding frequencies in Michigan during the warm summer months. Future prevention strategies aimed at reducing stress during lactation or isolating high-risk animals could be implemented to reduce herd-level shedding levels and avoid transmission of STEC to susceptible animals and people. IMPORTANCE: STEC shedding frequencies vary considerably across cattle herds in Michigan, and the shedding frequency of strains belonging to non-O157 serotypes far exceeds the shedding frequency of O157 strains, which is congruent with human infections in the state. Dairy cattle sampled at higher temperatures, in their first lactation, and early in the milk production stage were significantly more likely to shed STEC, which could be due to stress or a negative energy balance. Future studies should focus on the isolation of high-risk animals to decrease herd shedding levels and the potential for contamination of the food supply.

Association between genotypic diversity and biofilm production in group B Streptococcus.

BACKGROUND: Group B Streptococcus (GBS) is a leading cause of sepsis and meningitis and an important factor in premature and stillbirths. Biofilm production has been suggested to be important for GBS pathogenesis alongside many other elements, including phylogenetic lineage and virulence factors, such as pili and capsule type. A complete understanding of the confluence of these components, however, is lacking. To identify associations between biofilm phenotype, pilus profile and lineage, 293 strains from asymptomatic carriers, invasive disease cases, and bovine mastitis cases, were assessed for biofilm production using an in vitro assay. RESULTS: Multilocus sequence type (ST) profile, pilus island profile, and isolate source were associated with biofilm production. Strains from invasive disease cases and/or belonging to the ST-17 and ST-19 lineages were significantly more likely to form weak biofilms, whereas strains producing strong biofilms were recovered more frequently from individuals with asymptomatic colonization. CONCLUSIONS: These data suggest that biofilm production is a lineage-specific trait in GBS and may promote colonization of strains representing lineages other than STs 17 and 19. The findings herein also demonstrate that biofilms must be considered in the treatment of pregnant women, particularly for women with heavy GBS colonization.

Association and virulence gene expression vary among serotype III group B streptococcus isolates following exposure to decidual and lung epithelial cells.

Group B Streptococcus (GBS) causes severe disease in neonates, the elderly, and immunocompromised individuals. GBS species are highly diverse and can be classified by serotype and multilocus sequence typing. Sequence type 17 (ST-17) strains cause invasive neonatal disease more frequently than strains of other STs. Attachment and invasion of host cells are key steps in GBS pathogenesis. We investigated whether four serotype III strains representing ST-17 (two strains), ST-19, and ST-23 differ in their abilities to attach to and invade both decidual cells and lung epithelial cells. Virulence gene expression following host cell association and exposure to amnion cells was also tested. The ST-17 strains differed in their abilities to attach to and invade decidual cells, whereas there were no differences with lung epithelial cells. The ST-19 and ST-23 strains, however, attached to and invaded decidual cells less than both ST-17 strains. Although the ST-23 strain attached to lung epithelial cells better than ST-17 and -19 strains, none of the strains effectively invaded the lung epithelial cells. Notably, the association with host cells resulted in the differential expression of several virulence genes relative to basal expression levels. Similar expression patterns of some genes were observed regardless of cell type used. Collectively, these results show that GBS strains differ in their abilities to attach to distinct host cell types and express key virulence genes that are relevant to the disease process. Enhancing our understanding of pathogenic mechanisms could aid in the identification of novel therapeutic targets or vaccine candidates that could potentially decrease morbidity and mortality associated with neonatal infections.

Sepsis from the gut: the enteric habitat of bacteria that cause late-onset neonatal bloodstream infections.

BACKGROUND: Late-onset sepsis is a major problem in neonatology, but the habitat of the pathogens before bloodstream invasion occurs is not well established. METHODS: We examined prospectively collected stools from premature infants with sepsis to find pathogens that subsequently invaded their bloodstreams, and sought the same organisms in stools of infants without sepsis. Culture-based techniques were used to isolate stool bacteria that provisionally matched the bloodstream organisms, which were then genome sequenced to confirm or refute commonality. RESULTS: Of 11 children with late-onset neonatal bloodstream infections, 7 produced at least 1 stool that contained group B Streptococcus (GBS), Serratia marcescens, or Escherichia coli before their sepsis episode with provisionally matching organisms. Of 96 overlap comparison subjects without sepsis temporally associated with these cases, 4 were colonized with provisionally matching GBS or S. marcescens. Of 175 comparisons of stools from randomly selected infants without sepsis, 1 contained a GBS (this infant had also served as an overlap comparison subject and both specimens contained provisionally matching GBS). Genome sequencing confirmed common origin of provisionally matching fecal and blood isolates. The invasive E. coli were present in all presepticemic stools since birth, but gut colonization with GBS and S. marcescens occurred closer to time of bloodstream infection. CONCLUSIONS: The neonatal gut harbors sepsis-causing pathogens, but such organisms are not inevitable members of the normal microbiota. Surveillance microbiology, decolonization, and augmented hygiene might prevent dissemination of invasive bacteria between and within premature infants.

Diverse virulence gene content of Shiga toxin-producing Escherichia coli from finishing swine.

Shiga toxin-producing Escherichia coli (STEC) infections are a critical public health concern because they can cause severe clinical outcomes, such as hemolytic uremic syndrome, in humans. Determining the presence or absence of virulence genes is essential in assessing the potential pathogenicity of STEC strains. Currently, there is limited information about the virulence genes carried by swine STEC strains; therefore, this study was conducted to examine the presence and absence of 69 virulence genes in STEC strains recovered previously from finishing swine in a longitudinal study. A subset of STEC strains was analyzed by pulsed-field gel electrophoresis (PFGE) to examine their genetic relatedness. Swine STEC strains (n = 150) were analyzed by the use of a high-throughput real-time PCR array system, which included 69 virulence gene targets. Three major pathotypes consisted of 16 different combinations of virulence gene profiles, and serotypes were determined in the swine STEC strains. The majority of the swine STEC strains (n = 120) belonged to serotype O59:H21 and carried the same virulence gene profile, which consisted of 9 virulence genes: stx2e, iha, ecs1763, lpfAO113, estIa (STa), ehaA, paa, terE, and ureD. The eae, nleF, and nleH1-2 genes were detected in one swine STEC strain (O49:H21). Other genes encoding adhesins, including iha, were identified (n = 149). The PFGE results demonstrated that swine STEC strains from pigs raised in the same finishing barn were closely related. Our results revealed diverse virulence gene contents among the members of the swine STEC population and enhance understanding of the dynamics of transmission of STEC strains among pigs housed in the same barn.