Novel Chlamydia trachomatis Strains in Heterosexual Sex Partners,Indianapolis, Indiana,USA

Chlamydia trachomatis causes a high number of sexually transmitted infections worldwide, but reproducible and precise strain typing to link partners is lacking. We evaluated multilocus sequence typing (MLST) for this purpose by detecting sequence types (STs) concordant for the ompA genotype, a single-locus typing standard. We tested samples collected during April 2000–October 2003 from members of established heterosexual partnerships (dyads) in the Indianapolis, Indiana, USA, area who self-reported being coital partners within the previous 30 days. C. trachomatis DNA from 28 dyads was tested by MLST; sequences were aligned and analyzed for ST and phylogenetic relationships. MLST detected 9 C. trachomatis STs, 4 unique to Indianapolis; STs were identical within each dyad. Thirteen unique strains were identified; 9 (32%) dyads harbored novel recombinant strains that phylogenetically clustered with strains comprising the recombinants. The high rate of novel C. trachomatis recombinants identified supports the use of MLST for transmission and strain diversity studies among at-risk populations.     

Predicting Phenotype and Emerging Strains among Chlamydia trachomatis Infections

Chlamydia trachomatis is a global cause of blinding trachoma and sexually transmitted infections (STIs). We used comparative genomics of the family Chlamydiaceae to select conserved housekeeping genes for C. trachomatis multilocus sequencing, characterizing 19 reference and 68 clinical isolates from 6 continental/subcontinental regions. There were 44 sequence types (ST). Identical STs for STI isolates were recovered from different regions, whereas STs for trachoma isolates were restricted by continent. Twenty-nine of 52 alleles had nonuniform distributions of frequencies across regions (p<0.001). Phylogenetic analysis showed 3 disease clusters: invasive lymphogranuloma venereum strains, globally prevalent noninvasive STI strains (ompA genotypes D/Da, E, and F), and nonprevalent STI strains with a trachoma subcluster. Recombinant strains were observed among STI clusters. Single nucleotide polymorphisms (SNPs) were predictive of disease specificity. Multilocus and SNP typing can now be used to detect diverse and emerging C. trachomatis strains for epidemiologic and evolutionary studies of trachoma and STI populations worldwide.     

Phenotypic and molecular characterization of Streptococcus pneumoniae in pre-conjugate vaccine era: A Chinese hospital-based retrospective study

BackgroundStreptococcus pneumoniae (S. pneumoniae) is an important pathogen in causing global morbidity and mortality among children. This study aimed to determine phenotypic and molecular characteristics of S. pneumoniae causing infections in children under five years in China.MethodsA hospital-based retrospective study was conducted. All 537 S. pneumoniae isolates were tested for antimicrobial susceptibility by E-test method, molecular characteristics including resistance genes, virulence genes and serotypes by multiplex polymerase chain reaction (PCR) method, and sequence types (STs) by sequencing seven housekeeping genes. Minimum spanning tree and correspondence analysis were used to reveal the potential relationship between serotypes and STs.ResultsMost of S. pneumoniae isolates were resistant to erythromycin (93.9%) and tetracycline (86.4%), with the predominant resistance genes being erm(B) (92.6%) and tet(M) (95.5%). The prevalent serotypes were 19F, 6B, 19A, 23F and 14, the coverage rate of PCV13 was high in 85.8%, and the predominant STs were ST271, ST320, ST3173, ST81 and ST876. A significant correlation existed between STs and serotypes, with ST271/19F and ST320/19A as the most prevalent clones. Notably, ST271/19F and ST320/19A isolates were associated with resistance to specific antibiotics and carrying of mef(A/E), rlrA and sipA genes.ConclusionsOur findings suggest the introduction of PCV13 vaccine to Chinese children, and underscore the value of monitoring multiple characteristics to detect new epidemiologic trends and provide implications for the formulation of multivalent pneumococcal vaccines.     

Pneumococcal sequence type replacement among American Indian children: a comparison of pre- and routine-PCV7 eras

Background

Multi-locus sequence typing (MLST) of pneumococcal isolates collected during an efficacy trial of the 7-valent pneumococcal conjugate vaccine (PCV7) among Navajo and White Mountain Apache children from 1998 to 2000 showed a non-differential expansion of pre-existing sequence types (STs) and only one capsule-switching event in the PCV7-randomized communities. PCV7 was introduced as a routine infant vaccine in October 2000. We assessed variability in PCV7 effectiveness and mechanisms of ST replacement after prolonged routine PCV7 use.

Methods

We applied MLST to 267 non-vaccine type pneumococcal carriage and invasive disease isolates from Navajo and White Mountain Apache children from 2006 to 2008, and compared them to those from 1998 to 2000. Microarray was used to confirm capsule switching events.

Results

The primary mechanism of ST replacement among Navajo and White Mountain Apache children was expansion of existing STs, although introduction of new STs was an important secondary mechanism. ST199, a majority being serotype 19A, was the most common ST in both eras. Only ST193 (serotype 21) was preferentially expanding in the PCV7 era. Three examples of capsule switching were identified. No variability in vaccine effectiveness by ST was observed.

Conclusion

We did not observe an influence of ST on PCV7 serotype-specific effectiveness, although some STs may be favored in replacement.     

Carriage of Haemophilus influenzae in the oropharynx of young children and molecular epidemiology of the isolates after fifteen years of H. influenzae type b vaccination in Italy

BackgroundHaemophilus influenzae is an important pathogen able to cause a wide spectrum of diseases in children. Colonization of the upper respiratory tract is a risk factor for developing disease. This study aimed to investigate the oropharyngeal carriage rate of H. influenzae in young children in two Italian cities, 15 years after H. influenzae type b (Hib) vaccination was introduced. Antibiotic resistant traits and genotypes of the colonizing H. influenzae isolates were investigated.MethodsOropharyngeal swabs were obtained from 717 healthy children aged <6 years (June 2012–July 2013). Potential risk factors for H. influenzae colonization were investigated. H. influenzae isolates from carriage were characterized by PCR capsular typing, ampicillin susceptibility testing, resistance-associated gene sequencing and multilocus sequence typing (MLST). For comparison purposes, 38 non-typeable H. influenzae (NTHi) isolates from invasive disease were genotyped by MLST.ResultsThe overall H. influenzae carriage rate was 14.1% (101/717). Age, study site, presence of young siblings, and complete Hib vaccination status were independently associated with colonization. Of 101 isolates, 98 were NTHi, 2 were type e and 1 was type f. The overall ampicillin resistance rate was 15.8% (16/101). Resistance was mediated by TEM-1 β-lactamase production in half of isolates (n = 8) or modifications in penicillin-binding protein (PBP) 3 in the other half (n = 8). Several substitutions were discovered in PBP3 including the Asn526Lys change. Seventy-six different STs were identified among 98 NTHi isolates from carriage, with only 4 STs (ST12, ST57, ST238, ST1238) encompassing ≥3 isolates. Comparison of carriage and disease isolates found that several STs were shared between the two sources, although none of the major disease-associated STs were observed in carriage isolates.ConclusionsNTHi is the predominant serotype in carriage. The importance of monitoring both NTHi colonization rate and circulating genotypes should be emphasized in the era of the Hib conjugate vaccines.     

The clinical characteristics and genotype distribution of Chlamydia trachomatis infection in infants less than six months of age hospitalized with pneumonia

BackgroundChlamydia trachomatis is a common sexually-transmitted bacterial pathogen. As no routine screening is performed during pregnancy, neonates and infants are at high risk for C. trachomatis infection. The objective of this study was to investigate the morbidity, clinical characteristics and genotype distribution of C. trachomatis pneumonia in infants less than six months of age.MethodsClinical manifestations and laboratory results were recorded. Respiratory sputum specimens were tested using RT-PCR targeting C. trachomatis cryptic plasmid. Simultaneously, respiratory virus antigens were detected by direct immunofluorescence and bacterial pathogens were examined by culture in all sputum samples. Positive C. trachomatis samples were further genotyped using a multiplex PCR reverse line blot assay. The relationship between genotype and pneumonia severity was explored.ResultsOf 1408 infants, 101 (7.2%) were infected with C. trachomatis. Sixteen of 101 (15.8%) were assessed as severe pneumonia. These severe cases had a higher proportion of viral co-infection (37.5%) compared to mild pneumonia cases (9.4%, P < 0.05).Infants with tachypnea (OR 9.2) and wheezing (OR 3.5) were more likely to be classified as severe pneumonia (P < 0.05). Amongst 66 C. trachomatis specimens for which a genotyping result was available, seven genotypes were detected, and 39.4% of these specimens contained two or three genotypes. Overall, genotype E (48.5%) was the most frequent, followed by genotype F (42.4%), J (31.8%), D (12.1%), K (10.6%), G (4.5%) and H (3.0%). There were no significant correlations of particular genotypes with severity of disease, although there was a weak indication that more severe pneumonia might be associated with having certain mixed genotypes of C. trachomatis.ConclusionsThe prevalence of C. trachomatis in the population of young hospitalized infants with pneumonia in Shenzhen was very high. The relationship between genotype distribution and severity of pneumonia was not clear based on this study due to small sample size. Further in-depth investigation correlating genotype and disease severity based on a larger population is needed.     

Performance of urine leukocyte esterase in asymptomatic male youth: Another look with nucleic acid amplification testing as the gold standard for Chlamydia detection

PurposeTo re-evaluate the sensitivity and specificity of leukocyte esterase (LE) for screening adolescent and young adult males for Chlamydia trachomatis using a nucleic acid amplification test (NAAT) as the gold standard.MethodsThis study was conducted at two Massachusetts Department of Youth Services sites and one Job Corps site. Recently admitted asymptomatic sexually active male youth aged 14 to 25 years (mean 16.6 years) were recruited between January 2001 and July 2003 (N = 1008). Participants provided first part voided urine specimens for testing with LE and Chlamydia NAAT. The sensitivity, specificity, and positive and negative predictive value of urine LE for identification of Chlamydia infection were determined using NAAT as the gold standard.ResultsFifty-seven (5.7%) participants were infected with Chlamydia as defined by a positive NAAT. Defining trace + as the LE cut point resulted in sensitivity and specificity of 57.9% and 78.3%, respectively. Defining 1+ as the cut point resulted in sensitivity and specificity of 47.4% and 96.1%, respectively.ConclusionsUrine leukocyte esterase is a moderately sensitive method to screen for Chlamydia. Nevertheless, a substantial proportion of infections are not detected with LE screening. When feasible, urine NAAT provides a much more sensitive and equally noninvasive method of detecting Chlamydia. However, if LE is used as an initial screen followed by NAAT confirmation of LE positive samples, we recommend using trace LE as the cut point for positive results.     

Characterization of protective immune responses promoted by human antigen targets in a urogenital Chlamydia trachomatis mouse model

A vaccine against genital tract infections caused by Chlamydia trachomatis is urgently needed. We have previously identified a number of immunodominant human T- and/or B-cell antigen targets in patients with a C. trachomatis infection. Herein we use a urogenital C. trachomatis mouse model to investigate the protective efficacy of these antigens. C3H/HeN mice were immunized with recombinant antigens formulated in the adjuvant CAF01. Immunity post vaccination was analyzed and the protective efficacy against vaginal challenge with C. trachomatis was monitored by vaginal swabbing. All antigens elicited a significant immune response when administered in CAF01 but the balance between CMI and humoral responses differed markedly for the different antigens. Six (CT443, CT043, CT858, CT610, CT004 and CT681) antigens were found to be protective. We demonstrated by T-cell depletion studies that the protection promoted by the two antigens CT043 and CT004 was mediated by CD4⁺ T-cells. Both antigens are frequently recognized by T-cells during a natural Chlamydia infection in humans and if included in a future multi-component Chlamydia vaccine probably would operate mainly through the induction of a CMI response.     

Multilocus sequence typing (MLST) of leptospiral strains isolated from two geographic locations of Tamil Nadu,India

Here the rodent carrier status for the transmission of human leptospirosis in Tiruchirappalli, district, Tamil Nadu, India was assessed. The predominantly circulating leptospiral STs were recognized by multilocus sequence typing (MLST). A total of 113 rodents were trapped from different provinces of the Tiruchirappalli district. The most prevalent rodent was Bandicota bengalensis (37.2%), and of the total, 52.2% (n = 59) rodents were found to be positive for leptospiral 16S rRNA. These results were validated with a leptospiral culture positivity of 45.8% (n = 27). Three isolates from Chennai (2 rodents and 1 human) and 1 human isolate from Tiruchirappalli were included to understand the spatial variations and to track the source of human leptospirosis. The serogroup, serovar, and species level identification of all 31 isolates identified 28 to be Leptospira borgpetersenii serovar Javanica and three as Leptospira interrogans serovar Autumnalis. MLST analysis defined all isolates to the existing ST profiles (ST145 and ST27) with the exception of 6 L. borgpetersenii (ST DR) isolates that showed variations in the sucA and pfkB loci. The DR ST was locally confined to Chatram province of Tiruchirappalli suggesting an epidemiological link. The predominant STs, ST145 and ST-DR form a group, indicating the presence of original strain that subsequently diverged evolutionarily into two STs. The variations between L. borgpetersenii in sucA and pfkB loci may be an indication that evolutionary changes transpired in Tiruchirappalli.     

纹带棒状杆菌多位点序列分型及应用

目的 建立纹带棒状杆菌的多位点序列分型（MLST）方法,探讨临床分离纹带棒状杆菌的种群结构和遗传进化关系。方法 筛选出7个管家基因（gyrA、gyrB、hsp65、sodA、secA1、rpoB、16S rRNA）,设计引物并进行PCR扩增和测序,测序所得序列通过SeqMan软件进行拼接。采用DnaSP 5.10.01软件、Splits tree 4.14.2软件对管家基因的多样性及基因重组特征进行评价;采用MEGA 7.0.14软件基于序列型别（ST）采用M-L法构建系统发育树,采用BioNumerics软件基于ST特征值构建最小生成树,并用eBURST软件分析ST间遗传进化关系。结果 所选的7个位点在所有试验菌株中均获得了预期的扩增产物;Splits tree表明所有纹带棒状杆菌的聚类一致,提示基因重组是推动纹带棒状杆菌进化的潜在动力;MLST将344株纹带棒状杆菌分成72个STs,85.7%的菌株形成克隆复合体（CC）结构,CC19形成了优势克隆复合体,但包含菌株数最多的ST为该克隆复合体中的ST16。ST具有一定的地域聚集性且与分离年份具有一定的相关性。结论 我国纹带棒状杆菌呈现高度的遗传多样性,CC19为优势克隆复合体。本研究建立的MLST分型方案可用于纹带棒状杆菌的分型,但尚需优化改进。     

Outer membrane proteins preferentially load MHC class II peptides: Implications for a Chlamydia trachomatis T cell vaccine

CD4 T cell immune responses such as interferon-γ and tumor necrosis factor-α secretion are necessary for Chlamydia immunity. We used an immunoproteomic approach in which Chlamydia trachomatis and Chlamydia muridarum-derived peptides presented by MHC class II molecules on the surface of infected dendritic cells (DCs) were identified by tandem mass spectrometry using bone marrow derived DCs (BMDCs) from mice of different MHC background. We first compared the C. muridarum immunoproteome in C3H mice to that previously identified in C57BL/6 mice. Fourteen MHC class II binding peptides from 11 Chlamydia proteins were identified from C3H infected BMDCs. Two C. muridarum proteins overlapped between C3H and C57B/6 mice and both were polymorphic membrane proteins (Pmps) which presented distinct class II binding peptides. Next we studied DCs from C57BL/6 mice infected with the human strain, C. trachomatis serovar D. Sixty MHC class II binding peptides derived from 27 C. trachomatis proteins were identified. Nine proteins were orthologous T cell antigens between C. trachomatis and C. muridarum and 2 of the nine were Pmps which generated MHC class II binding epitopes at distinct sequences within the proteins. As determined by antigen specific splenocyte responses outer membrane proteins PmpF, -G and -H and the major outer membrane protein (MOMP) were antigenic in mice previously infected with C. muridarum or C. trachomatis. Furthermore a recombinant protein vaccine consisting of the four Pmps (PmpEFGH) with MOMP formulated with a Th1 polarizing adjuvant significantly accelerated (p < 0.001) clearance in the C57BL/6 mice C. trachomatis transcervical infection model. We conclude that Chlamydia outer membrane proteins are important T cell antigens useful in the development of a C. trachomatis subunit vaccine.     

Chlamydia trachomatis control requires a vaccine

As the most common reported communicable disease in North America and Europe, Chlamydia trachomatis is the focus of concerted public health control efforts based on screening and treatment. Unexpectedly control efforts are accompanied by rising reinfection rates attributed in part to arresting the development of herd immunity. Shortening the duration of infection through the testing and treatment program is the root cause behind the arrested immunity hypothesis and because of this a vaccine will be essential to control efforts. Advances in Chlamydia vaccinomics have revealed the C. trachomatis antigens that can be used to constitute a subunit vaccine and a vaccine solution appears to be scientifically achievable. We propose that an accelerated C. trachomatis vaccine effort requires coordinated partnership among academic, public health and private sector players together with a commitment to C. trachomatis vaccine control as a global public health priority.     

Chlamydia trachomatis: Cell biology,immunology and vaccination

Chlamydia trachomatis is the causative agent of a highly prevalent sexually transmitted bacterial disease and is associated with a number of severe disease complications. Current therapy options are successful at treating disease, but patients are left without protective immunity and do not benefit the majority asymptomatic patients who do not seek treatment. As such, there is a clear need for a broad acting, protective vaccine that can prevent transmission and protect against symptomatic disease presentation. There are three key elements that underlie successful vaccine development: 1) Chlamydia biology and immune-evasion adaptations, 2) the correlates of protection that prevent disease in natural and experimental infection, 3) reflection upon the evidence provided by previous vaccine attempts. In this review, we give an overview of the unique intra-cellular biology of C. trachomatis and give insight into the dynamic combination of adaptations that allow Chlamydia to subvert host immunity and survive within the cell. We explore the current understanding of chlamydial immunity in animal models and in humans and characterise the key immune correlates of protection against infection. We discuss in detail the specific immune interactions involved in protection, with relevance placed on the CD4+ T lymphocyte and B lymphocyte responses that are key to pathogen clearance. Finally, we provide a timeline of C. trachomatis vaccine research to date and evaluate the successes and failures in development so far. With insight from these three key elements of research, we suggest potential solutions for chlamydial vaccine development and promising avenues for further exploration.     

Induction of protective immunity by vaccination against Chlamydia trachomatis using the major outer membrane protein adjuvanted with CpG oligodeoxynucleotide coupled to the nontoxic B subunit of cholera toxin

The present study was undertaken to test the efficacy of immunization with the native major outer membrane protein (nMOMP) of Chlamydia trachomatis mouse pneumonitis (MoPn) serovar in combination with a novel immunostimulatory adjuvant consisting of CpG oligodeoxynucleotide (ODN) linked to the nontoxic B subunit of cholera toxin (CTB-CpG) to elicit a protective immune response to C. trachomatis. High levels of Chlamydia-specific IgG antibodies were detected in the sera from BALB/c mice immunized intramuscularly and subcutaneously (i.m. + s.c.) with the nMOMP/CTB-CpG vaccine or with nMOMP adjuvanted with a mixture of CT and CpG ODN (CT + CpG). Further, these immunization schemes gave rise to significant T-cell-mediated Chlamydia-specific immune responses. No Chlamydia-specific humoral or cell-mediated immune responses were detected in the control mice vaccinated with ovalbumin together with either CTB-CpG or CT + CpG. Following an intranasal challenge with C. trachomatis the groups of mice immunized with nMOMP plus CTB-CpG, CT + CpG or live C. trachomatis were found to be protected based on their change in body weight and lung weight as well as number of inclusion forming unit recovered from the lungs, as compared with control groups immunized with ovalbumin plus either adjuvants. Interestingly, IFN-γ-producing CD4⁺, but not CD8⁺, T-cells showed a significant correlation with the outcomes of the challenge. In conclusion, nMOMP in combination with the novel adjuvant CTB-CpG elicited a significant antigen-specific antibody and cell-mediated immune responses as well as protection against a pulmonary challenge with C. trachomatis.     

Molecular characteristics of Streptococcus agalactiae in a mother-baby prospective cohort study: Implication for vaccine development and insights into vertical transmission

BackgroundStreptococcus agalactiae (GBS) is a leading cause of neonatal sepsis and meningitis in many countries. This study aimed to determine the molecular characteristics of GBS colonized in mothers and their infants so as to provide implication for vaccine strategies and confirm vertical transmission.MethodsA prospective cohort study was conducted to recruit 1815 mother-neonate pairs. All GBS isolates from pregnant women and her infants were tested for serotypes, multilocus sequence types and virulence genes. The relationship between multiple molecular characteristics of GBS isolates was tested by the correspondence analysis, and the agreement between mother-neonate paired data in molecular characteristics was analyzed using Kappa tests.ResultsThe predominant serotypes were III, Ia and V, and the most prevalent sequence types (STs) were ST19, ST17, ST10, and ST12. All isolates carried at least one pilus island (PI). The most common combination of PIs was PI-2b alone, followed by PI-1+PI-2a and PI-2a alone, and the most prevalent alpha-like protein (alp) genes were rib, epsilon and alphaC. Moreover, a strong relationship was noted between STs, serotypes, alp genes and PIs, including ST17 associated with serotype-III/rib/PI-2b, ST19 with serotype-III/rib/PI-1+PI-2a, and ST485 with serotype-Ia/epsilon/PI-2b. The rate of GBS vertical transmission was 14.1%, and the kappa test revealed good agreement in multiple molecular characteristics among GBS-positive mother-neonate pairs. Notably, the switching of molecular characteristics was found during vertical transmission.ConclusionsOur findings underscore the value of monitoring multiple molecular characteristics so as to provide implication for multivalent strategies and gain insights into GBS vertical transmission and vertical characteristic switching.     

Clonal complexes 104, 109 and 113 playing a major role in the dissemination of OXA-carbapenemase-producing Acinetobacter baumannii in Southeast Brazil

Carbapenem resistance among Acinetobacter baumannii strains isolated from clinical settings in Brazil has increased dramatically in the last 10 years due to the emergence and dissemination of OXA-type carbapenemase encoding genes. This study aimed to characterize the presence of carbapenem-hydrolyzing class D β-lactamases (CHDL)-encoding genes and clonal complexes playing a major role in the dissemination of OXA-carbapenemase-producing A. baumannii in Southeast Brazil.A total of 74 A. baumannii strains isolated from patients admitted to 4 hospitals in Southeast Brazil were analyzed. Molecular characterization of strains revealed that 67 strains carried bla_OXA-23 (72%), bla_OXA-143 (25%) or both genes (3%). PFGE analysis identified 12 PFGE clusters, grouping 26 pulsotypes. Two PFGE clusters were predominant, comprising more than 66% of OXA-producing A. baumannii isolates. Among 23 representative strains characterized by MLST-UO (Multilocus Sequence Typing Scheme – University of Oxford, http://pubmlst.org/abaumannii/), 14 different STs were identified, of which six were confirmed as novel sequence types (designated as STs 402–407). Most of these isolates belonged to clonal complexes CC104,CC109 or CC113, whereas three STs were singletons (ST339, 403 and 407).In conclusion, the presence of bla_OXA-23- and bla_OXA-143-like genes was not related to specific ST/CC, suggesting that the dissemination of OXA-carbapenemase-encoding genes may involve different STs, in which the spread of OXA-23-like is most likely due to mobile elements (i.e., plasmids). In this regard, CC104, CC109 and CC113 played a major role as predominant CDHL-carrying clones, instead of CC92, which was not identified.     

A T cell epitope-based vaccine protects against chlamydial infection in HLA-DR4 transgenic mice

Vaccination with recombinant chlamydial protease-like activity factor (rCPAF) has been shown to provide robust protection against genital Chlamydia infection. Adoptive transfer of IFN-γ competent CPAF-specific CD4⁺ T cells was sufficient to induce early resolution of chlamydial infection and reduction of subsequent pathology in recipient IFN-γ-deficient mice indicating the importance of IFN-γ secreting CD4⁺ T cells in host defense against Chlamydia. In this study, we identify CD4⁺ T cell reactive CPAF epitopes and characterize the activation of epitope-specific CD4⁺ T cells following antigen immunization or Chlamydia challenge. Using the HLA-DR4 (HLA-DRB1*0401) transgenic mouse for screening overlapping peptides that induced T cell IFN-γ production, we identified at least 5 CPAF T cell epitopes presented by the HLA-DR4 complex. Immunization of HLA-DR4 transgenic mice with a rCPAFep fusion protein containing these 5 epitopes induced a robust cell-mediated immune response and significantly accelerated the resolution of genital and pulmonary Chlamydia infection. rCPAFep vaccination induced CPAF-specific CD4⁺ T cells in the spleen were detected using HLA-DR4/CPAF-epitope tetramers. Additionally, CPAF-specific CD4⁺ clones could be detected in the mouse spleen following Chlamydia muridarum and a human Chlamydia trachomatis strain challenge using these novel tetramers. These results provide the first direct evidence that a novel CPAF epitope vaccine can provide protection and that HLA-DR4/CPAF-epitope tetramers can detect CPAF epitope-specific CD4⁺ T cells in HLA-DR4 mice following C. muridarum or C. trachomatis infection. Such tetramers could be a useful tool for monitoring CD4⁺ T cells in immunity to Chlamydia infection and in developing epitope-based human vaccines using the murine model.     

A comparison of the effects of a chlamydial vaccine administered during or after a C. muridarum urogenital infection of female mice

Research into an efficacious Chlamydia trachomatis vaccine is ongoing, however, there has been no examination into the timing of vaccine administration to either asymptomatically or previously infected individuals. Using the female Chlamydia muridarum genital tract mouse model, we examined this aspect of vaccine development. Our results show timing of vaccination affected the production of systemic antibodies, but had minimal effects on mucosal antibody production. Vaccination during an active infection or after a resolved infection did not provide protection against re-exposure to Chlamydia, and did not exacerbate the development of pathological sequelae in infected mice. This demonstrates that vaccination may not be protective in individuals who are seropositive for an acute or previous chlamydial infection.     

Immunogenicity of a vaccine formulated with the Chlamydia trachomatis serovar F, native major outer membrane protein in a nonhuman primate model

To determine the ability of a vaccine formulated with the genital Chlamydia trachomatis, serovar F, native major outer membrane protein (Ct-F-nMOMP), to induce systemic and mucosal immune responses, rhesus macaques (Macaca mulatta) were immunized three times by the intramuscular (i.m.) and subcutaneous (s.c.) routes using CpG-2395 and Montanide ISA 720 VG, as adjuvants. As controls, another group of M. mulatta was immunized with ovalbumin instead of Ct-F-nMOMP using the same formulation and routes. High levels of Chlamydia-specific IgG and IgA antibodies were detected in plasma, vaginal washes, tears, saliva, and stools from the Ct-F-nMOMP immunized animals. Also, high neutralizing antibody titers were detected in the plasma from these animals. Monkeys immunized with ovalbumin had no detectable Chlamydia-specific antibodies. Furthermore, as measured by a lymphoproliferative assay, significant Chlamydia-specific cell-mediated immune responses were detected in the peripheral blood mononuclear cells (PBMC) from the rhesus macaques vaccinated with Ct-F-nMOMP when compared with the animals immunized with ovalbumin. In addition, the levels of two Th1 cytokines, IFN-γ and TNF-α, were significantly higher in the animals immunized with Ct-F-nMOMP when compared with those from the monkeys immunized with ovalbumin. To our knowledge, this is the first time that mucosal and systemic immune responses have been investigated in a nonhuman primate model using a subunit vaccine from a human genital C. trachomatis serovar.     

Genetic analysis of human clinical isolates of Lactococcus garvieae: Relatedness with isolates from foods

Lactococcus garvieae is a Gram-positive bacterium well-known as an important pathogen in aquaculture, and it is also a human pathogen of increasing clinical significance. Forty-three human L. garvieae isolates from clinical specimens were characterized by Multilocus Sequence Typing (MLST). Twenty-six different sequence types (STs) were identified among the human isolates, of which 20 were novel STs. Most human isolates clustered into four clonal complexes, with a predominance of CC3. Within CC3, ST10 was the most common genotype, indicating the existence of a circulating genetic lineage among the human isolates analyzed. The four CCs also grouped L. garvieae strains isolated from meat, dairy and fish, indicating a genetic overlap between isolates from human and these foods. Genetic relatedness among human and food L. garvieae isolates was confirmed by phylogenetic analysis based on the concatenated sequences of the seven MLST genes. These results represent the first evidence of genetic relatedness between isolates of L. garvieae of human and those isolated meat, milk and dairy products and suggest that, in addition to fish and seafood, these foods might represent important sources of human L. garvieae infections.