Effect of alpha-oligosaccharide phenotype of Neisseria gonorrhoeae strain MS11 on invasion of Chang conjunctival, HEC-1-B endometrial, and ME-180 cervical cells

The genes encoding the glycosyltransferases responsible for the addition of the five sugars in the alpha-oligosaccharide (alpha-OS) moiety of lipooligosaccharide (LOS) have been identified. Disruption of these glycosyltransferase genes singly or in combination results in corresponding truncations in LOS. In the present work we show that sequential deletion of the terminal four sugar residues of gonococcal alpha-OS had no discernible effect on the invasion of human conjunctival, endometrial, and cervical cell lines. However, deletion of the proximal glucose, which resulted in the complete deletion of alpha-OS, significantly impaired invasion of the gonococci into all three cell lines. The effect of deleting alpha-OS on invasion was independent of and additive to the known invasion-promoting factor OpaA. These data suggest that the proximal glucose residue of the alpha-OS chain of LOS is required for efficient invasion of gonococci into host mucosa.     

Identification of Regions of the Chromosome of Neisseria meningitidis and Neisseria gonorrhoeae Which Are Specific to the Pathogenic Neisseria Species

Neisseria meningitidis and Neisseria gonorrhoeae give rise to dramatically different diseases. Their interactions with the host, however, do share common characteristics: they are both human pathogens which do not survive in the environment and which colonize and invade mucosa at their port of entry. It is therefore likely that they have common properties that might not be found in nonpathogenic bacteria belonging to the same genetically related group, such as Neisseria lactamica. Their common properties may be determined by chromosomal regions found only in the pathogenic Neisseria species. To address this issue, we used a previously described technique (C. R. Tinsley and X. Nassif, Proc. Natl. Acad. Sci. USA 93:11109-11114, 1996) to identify sequences of DNA specific for pathogenic neisseriae and not found in N. lactamica. Sequences present in N. lactamica were physically subtracted from the N. meningitidis Z2491 sequence and also from the N. gonorrhoeae FA1090 sequence. The clones obtained from each subtraction were tested by Southern blotting for their reactivity with the three species, and only those which reacted with both N. meningitidis and N. gonorrhoeae (i.e., not specific to either one of the pathogens) were further investigated. In a first step, these clones were mapped onto the chromosomes of both N. meningitidis and N. gonorrhoeae. The majority of the clones were arranged in clusters extending up to 10 kb, suggesting the presence of chromosomal regions common to N. meningitidis and N. gonorrhoeae which distinguish these pathogens from the commensal N. lactamica. The sequences surrounding these clones were determined from the N. meningitidis genome-sequencing project. Several clones corresponded to previously described factors required for colonization and survival at the port of entry, such as immunoglobulin A protease and PilC. Others were homologous to virulence-associated proteins in other bacteria, demonstrating that the subtractive clones are capable of pinpointing chromosomal regions shared by N. meningitidis and N. gonorrhoeae which are involved in common aspects of the host interaction of both pathogens.     

Ultrastructure of Pili and Annular Structures on the Cell Wall Surface of Neisseria meningitidis

In a survey of negatively stained preparations of the prototype strains of Neisseria meningitidis, pili were detected on three strains. However, these pili were detected on fewer than 5% of cells in populations of these three strains. Those individual cells with pili were seldom observed to contain more than two or three pili per cell. In contrast, nearly all cells of the nonprototype group B strain ATCC 13090 had numerous pili on their surfaces. When viewed in frozen-etched replicas, a few pili were observed lying on the cell surface of this latter strain. An annular structure was also found in frozen-etched replicas. This structure usually consisted of a series of concentric rings that were always found on the flat side of these bean-shaped cells. It is concluded that such structures represent a differentiated portion of the cell wall, which is involved in cross-wall formation during synthesis of the cell septum in dividing cells.     

Nucleotide sequence homology between the immunoglobulin A1 protease genes of Neisseria gonorrhoeae, Neisseria meningitidis, and Haemophilus influenzae.

Isolated DNA fragments encoding the immunoglobulin A1 (IgA1) protease of Neisseria gonorrhoeae were used as hybridization probes to search for homologous sequences in whole cell DNA from Neisseria meningitidis and Haemophilus influenzae. Significant homology was detected. That the detected homology represented IgA1 protease-specific sequences was confirmed by the cloning of these sequences in Escherichia coli HB101 and demonstrating the expression of IgA1 protease by these transformed cells. Molecular probing of commensal Neisseria and Haemophilus species, which do not elaborate IgA1 protease activity, revealed that they were devoid of sequence homology with the cloned IgA1 protease gene DNA.     

Cell envelope of Neisseria gonorrhoeae: relationship between autolysis in buffer and the hydrolysis of peptidoglycan.

Neisseria gonorrhoeae readily underwent autolysis when suspended in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) buffer at alkaline pH values. Autolysis was inhibited by the addition of Mg2+ or other divalent cations. Autolysis was also suppressed at acid pH (pH 6.0). Suspension of cells in buffer was accompanied by the hydrolysis of peptidoglycan. The rate of peptidoglycan hydrolysis in HEPES buffer was maximal at pH 8.5 and was similar in the presence or absence of Mg2+. Therefore, divalent cation stabilization against autolysis is not mediated by inhibition of peptidoglycan hydrolysis. Peptidoglycan hydrolysis occurred in HEPES buffer (pH 6.0), but at a rate that was 50% of the maximum. Incubation of cells with chloramphenicol or rifampin before suspension in HEPES buffer (pH 8.5) partially prevented autolysis; under these conditions, peptidoglycan hydrolysis still occurred, but at a reduced rate. Old and new peptidoglycans were hydrolyzed at similar rates. Peptidoglycan hydrolysis results in solubilization of both the peptide and glycan moieties.     

Identification, localization, and distribution of the PilT protein in Neisseria gonorrhoeae.

A monoclonal antibody (MAb) directed against a highly conserved protein of Neisseria gonorrhoeae with a molecular size of 40 kDa was isolated and characterized. The protein antigen detected by this MAb was detected by enzyme-linked immunosorbent assay and immunoblotting in all strains of N. gonorrhoeae tested across a wide range of serovars. The 40-kDa protein was found to be expressed at relatively low levels and localized to both the cytosolic and cytoplasmic membrane fractions. Screening of a lambda gt11 expression library derived from gonococcal genomic DNA with the anti-40-kDa MAb and DNA sequence analysis suggested that the 40-kDa protein and the product of the gonococcal pilT gene were identical. Immunoblotting analysis of gonococcal mutants carrying defined mutations in the pilT gene confirmed that the 40-kDa protein was indeed PilT. The N-terminal sequence derived by microsequencing of the protein purified from gonococci led to the correction of the previously published pilT gene sequence. Sequencing of the pilT gene from three different strains revealed an extremely high degree of conservation at both the amino acid and DNA levels.     

Ability of the Digene Hybrid Capture II Test To Identify Chlamydia trachomatis and Neisseria gonorrhoeae in Cervical Specimens

The Digene Hybrid Capture II (HCII CT/GC) test is a combination test designed to detect Chlamydia trachomatis and Neisseria gonorrhoeae in a single specimen. It is a nucleic acid hybridization test which uses signal amplification to increase sensitivity. We compared its performance to that of culture on cervical specimens from 1,370 women. Direct fluorescent-antibody assay was used to resolve discrepant results for C. trachomatis. Samples were collected with a proprietary cervical brush or with endocervical swabs. The HCII CT/GC test proved to be sensitive and specific in detecting these organisms. Compared to N. gonorrhoeae culture, it had a sensitivity of 93% (87/94) and a specificity of 98.5% (1,244/1,263). Compared to C. trachomatis culture, the sensitivity was 97.7% (129/132) and specificity was 98.2% (1,216/1,238). Testing of some specimens with discrepant results by PCR suggested that the test would actually prove to be even more specific if it were compared to a nucleic acid amplification test (NAAT). The sensitivity of C. trachomatis culture was somewhat less, at 88.6% (117/132). The endocervical brush appeared to be better than Dacron swabs for collecting specimens. The HCII CT/GC test offers an attractive format that allows simultaneous detection of C. trachomatis and N. gonorrhoeae with a single specimen. An initial positive result is followed by repeat tests with probes to identify chlamydiae or gonococci. This test is more sensitive than C. trachomatis culture and is at least as sensitive as culture for gonococci. It deserves further evaluation and comparison with NAATs and may well offer an attractive alternative for diagnosis and screening of these infections.     

Role of endotoxin in the pathogenicity of Neisseria gonorrhoeae colonial types 1, 4 and 5 determined by chicken embryo model

The pathogenicity of Neisseria gonorrhoeae is a subject of considerable interest. It is believed that N. gonorrhoeae of colonial type 1 are pathogenic while those of type 4 are not. This is based on experimentation in human volunteers. The object of this study was to determine the reasons for the differences of susceptibility of chicken embryos to N. gonorrhoeae strains of colonial types 1, 4, 5 and 1R (a type-1 revertant from a non-pathogenic type 4 strain originally tested in human volunteers). Colonial types 1, 5, 1R and 4 caused mortality rates of 80, 70, 85 and 20% respectively. This variation in lethality appeared to depend upon the availability of free extra-cellular endotoxin and this was confirmed by chicken-embryo inoculation results and electronmicroscopy of normal and heated colonial types 1, 4 and 5. Similar results were obtained by inoculating purified endotoxins from these types into chicken embryos. The results of this study suggest that endotoxins play a major role in the pathogenicity of N. gonorrhoeae and that the variations in virulence of the colonial types depends on the stability of their cell walls.     

Development and Evaluation of a Magnetic Immunochromatographic Test To Detect Taenia solium,Which Causes Taeniasis and Neurocysticercosis in Humans

Taeniasis/cysticercosis caused by Taenia solium is a frequent parasitic infection of the human brain in most of the world. Rapid and simple screening tools to identify taeniasis and cysticercosis cases are needed for control programs, mostly to identify tapeworm carriers which are the source of infection and need to be treated, or as tools for point-of-care case detection or confirmation. These screening assays should be affordable, reliable, rapid, and easy to perform. Immunochromatographic tests meet these criteria. To demonstrate proof of principle, we developed and evaluated two magnetic immunochromatographic tests (MICTs) for detection of human Taenia solium taeniasis antibodies (ES33-MICT) and neurocysticercosis antibodies (T24-MICT). These assays detected stage-specific antibodies by using two recombinant proteins, rES33 for detection of taeniasis antibodies and rT24H for detection of cysticercosis antibodies. The sensitivity and specificity of the ES33-MICT to detect taeniasis infections were 94.5% and 96%, respectively, and those of the T24-MICT to detect cases of human cysticercosis with two or more viable brain cysts were 93.9% and 98.9%, respectively. These data provide proof of principle that the ES33- and T24-MICTs provide rapid and suitable methods to identify individuals with taeniasis and cysticercosis.Humans can be infected with both the adult worm and larval forms of the cestode Taenia solium, causing taeniasis and cysticercosis, respectively. Humans, who are the only definitive host for the adult tapeworm, develop taeniasis after eating raw or undercooked pork infested with the larval stages of the parasite. Both pigs and humans can develop cysticercosis by ingesting eggs passed in the feces of a tapeworm carrier. Consequently, cysticercosis can be acquired under any variety of cultural and socioeconomic conditions where there is close contact with a taeniasis carrier. Because tapeworm-infected humans are the only source of transmission of both human and pig cysticercosis, diagnosis and treatment of taeniasis are crucial for control and elimination of cysticercosis in both humans and pigs (18). Therefore, accurate and rapid diagnostic methods to identify tapeworm carriers are critical tools needed in cysticercosis/taeniasis control and elimination programs. Although diagnosis of porcine or human cysticercosis is not crucial in the context of control programs, the availability of rapid diagnosis for human cysticercosis would be useful for estimating the burden of disease and for determining seroprevalence rates in pigs.Diagnosis of taeniasis usually is established by microscopic observation of T. solium eggs in stool specimens, but this method is insensitive and cannot differentiate between T. solium and Taenia saginata ova. Other methods to detect taeniasis include coproantigen detection methods, which are more sensitive than microscopy, and serodiagnosis of taeniasis by the use of excretory-secretory proteins in an enzyme-linked immunoelectrotransfer blot (EITB) (1, 14, 23). Two specific T. solium excretory-secretory (TSES) proteins, ES33 and ES38, were identified for specific taeniasis serodiagnosis. Recombinant forms of ES33 and ES38 (rES33 and rES38) have been evaluated, and the diagnostic performances of these two proteins were determined to be comparable (11).The gold standard for serological identification of cysticercosis is the EITB, which relies on antibody reactivity with seven diagnostic lentil lectin purified glycoproteins (LLGP) (20). Recombinant or synthetic peptide forms of these proteins are now available (7-9), and after comparing the performances of these diagnostic proteins, rT24H (which corresponds to a 24,000-Da protein of the LLGP extract) (9) was identified as the recombinant protein with the best sensitivity and specificity for detecting neurocysticercosis cases (11).The gold standard tests described above are not field friendly, not widely available, and do not exist in point-of-care formats. In contrast, lateral flow immunochromatographic tests, can be used at the point of care and in settings with minimal infrastructure (5, 21). Lateral flow methods have been used for several decades for clinical and veterinary diagnosis (16). In conventional lateral flow assays, reactions are detected visually, generating a qualitative result, or with a detection device that measures reflectance, contrast, color change, or fluorescence (16). Several studies have demonstrated that magnetic particle-labeled detection systems have the potential to improve lateral flow assay sensitivity and to provide a means of quantification (12, 17, 25). In this study, we describe the development and evaluation of two magnetic immunochromatographic tests (MICTs) for serologic detection of taeniasis (based on rES33 antigen) and cysticercosis (based on the rT24H antigen).     

Neisseria gonorrhoeae porin P1.B induces endosome exocytosis and a redistribution of Lamp1 to the plasma membrane

The immunoglobulin A (IgA) protease secreted by pathogenic Neisseria spp. cleaves Lamp1, thereby altering lysosomes in a cell and promoting bacterial intracellular survival. We sought to determine how the IgA protease gains access to cellular Lamp1 in order to better understand the role of this cleavage event in bacterial infection. In a previous report, we demonstrated that the pilus-induced Ca(2+) transient triggers lysosome exocytosis in human epithelial cells. This, in turn, increases the level of Lamp1 at the plasma membrane, where it can be cleaved by IgA protease. Here, we show that porin also induces a Ca(2+) flux in epithelial cells. This transient is similar in nature to that observed in phagocytes exposed to porin. In contrast to the pilus-induced Ca(2+) transient, the porin-induced event does not trigger lysosome exocytosis. Instead, it stimulates exocytosis of early and late endosomes and increases Lamp1 on the cell surface. These results indicate that Neisseria pili and porin perturb Lamp1 trafficking in epithelial cells by triggering separate and distinct Ca(2+)-dependent exocytic events, bringing Lamp1 to the cell surface, where it can be cleaved by IgA protease.     

Mutation Pattern in the Genome of Neisseria gonorrhoeae and Its Association with Multidrug-resistant Isolates from Delhi,India

Mutations in PenA, PorB, MtrE and MtrR genes responsible for antimicrobial resistance were checked in 27 drug-resistant clinical isolates of Neisseria gonorrhoeae (NG). Phenotype PIB (88.88%) and mutation at G120 and A121 positions of porB were recurrent. N122K, a novel mutation, was observed in PorB in three resistant isolates. Substitution H105Y in MtrR was widespread (37% of clinical isolates). The presence of a novel mutation (L33V), along with G45D mutation in MtrR, was associated with less-resistant isolates, in contrast to isolates with G45D mutation alone. African-type penicillinase-producing NG plasmid was observed most frequently (17/27) in penicillin-resistant isolates.     

Molecular characterization of the argJ mutation in Neisseria gonorrhoeae strains with requirements for arginine, hypoxanthine, and uracil.

Arginine auxotrophs are commonly encountered among clinical isolates of Neisseria gonorrhoeae. Arginine auxotrophs which also require hypoxanthine and uracil (AHU strains) compose a unique set of strains that are highly homogeneous and are believed to be clonally derived. The Arg- phenotype of these strains is due to a lesion in the argJ gene encoding ornithine acetyltransferase. We have cloned the mutant argJ gene from an AHU strain and compared the sequence of this gene to the wild-type argJ gene. The mutant gene contained a 3-bp deletion within a repetitive region of the argJ gene. This mutation was restored to the wild-type sequence in a naturally occurring Arg+ revertant of the AHU strain. This deletion was detected in a wide variety of other AHU strains but not in other ArgJ- strains or in ArgJ+ strains, supporting the theory that AHU strains are clonally derived.     

Emerging antimicrobial resistance, plasmid profile and pulsed-field gel electrophoresis pattern of the endonuclease-digested genomic DNA of Neisseria gonorrhoeae

Resistant gonococci are very prevalent in many countries, particularly in Asia. This study was conducted to determine the trend of resistance, the effect of decreasing the ciprofloxacin susceptibilities of gonococci on the prevalence of penicillinase-producing N. gonorrhoeae (PPNG), and to compare the epidemiology of strains with the previous studies. A total of 602 strains of gonococci were isolated from prostitutes in 1997-1999. Antimicrobial susceptibility was tested by NCCLS disk diffusion and agar dilution methods. For epidemiologic analysis, plasmid analysis and pulsed-field gel electrophoresis (PFGE) were performed. The proportion of PPNG remained high (79%), and the strains with decreased susceptibility to ciprofloxacin increased significantly from 67% in 1997 to 84% in 1999. Compared to our previous study, the PFGE patterns were similar, while the proportion of strain with the 3.2-MDa plasmid markedly decreased. In conclusion, a rapid increase in ciprofloxacin-nonsusceptible strains may suggest difficulties in the treatment of gonococcal infections in the near future with the drug. The recent decrease of PPNG with the 3.2-MDa plasmid may suggest that there is an epidemiological change in gonococcal infections, and the prevalence of related PFGE patterns suggests the dissemination of a few clones among the high risk populations.     

Comparison of microscopy, culture and enzyme immunoassay (Gonozyme) for the detection of Neisseria gonorrhoeae in urogenital specimens

Urogenital specimens of male patients and female prostitutes were examined for gonorrhoea in a gonococcal antigen enzyme immunoassay (Gonozyme), by microscopic examination of stained smears and by bacterial culture. Out of 18 male patients, 14 showed positive reactions (all 14 by Gonozyme and by microscopy, but only eight by culture also). The sensitivity and specificity of Gonozyme was 100% in reference to microscopy. The predictive value for a positive test and for a negative test was 100%. The sensitivity of Gonozyme in reference to culture was also 100%, but the specificity was only 40%, because of the low yield of positive cultures. The predictive value for a positive test was 57% and for a negative test 100%. Out of 189 female prostitutes, 41 (22%) had a positive reaction in at least one test (Gonozyme, microscopy and culture were positive in 10; Gonozyme and culture in three; Gonozyme and microscopy in 14; Gonozyme alone in 11; culture alone in three). The sensitivity of Gonozyme was 100% and specificity 92% in reference to microscopy. The predictive value for a positive test was 63% and for a negative test 100%. In reference to culture, the sensitivity was 81% and specificity 86%. The predictive value for a positive test was 34% and for a negative test 98%. In prostitutes, the rate of asymptomatic infections was 14%, if one assumed that all Gonozyme-positive results were truly positive. Gonozyme proved to be the most sensitive method for screening female patients. To discriminate possibly false positive reactions, Gonozyme-positive specimens should be corroborated, preferably by bacterial cultivation.     

Allelic and haplotypic diversity of HLA-A, -B, -C, -DRB1, and -DQB1 genes in the Korean population

High-resolution human leukocyte antigen (HLA) typing exposes the unique patterns of HLA allele and haplotype frequencies in each population. In this study, HLA-A, -B, -C, -DRB1, and -DQB1 genotypes were analyzed in 485 apparently unrelated healthy Korean individuals. A total of 20 HLA-A, 43 HLA-B, 21 HLA-C, 31 HLA-DRB1, and 14 HLA-DQB1 alleles were identified. Eleven alleles (A*0201, A*1101, A*2402, A*3303, B*1501, Cw*0102, Cw*0302, Cw*0303, DQB1*0301, DQB1*0302, and DQB1*0303) were found in more than 10% of the population. In each serologic group, a maximum of three alleles were found with several exceptions (A2, B62, DR4, DR14, and DQ6). In each serologic group exhibiting multiple alleles, two major alleles were present at 62-96% (i.e. A*0201 and A*0206 comprise 85% of A2-positive alleles). Multiple-locus haplotypes estimated by the maximum likelihood method revealed 51 A-C, 43 C-B, 52 B-DRB1, 34 DRB1-DQB1, 48 A-C-B, 42 C-B-DRB1, 46 B-DRB1-DQB1, and 30 A-C-B-DRB1-DQB1 haplotypes with frequencies of more than 0.5%. In spite of their high polymorphism in B and DRB1, identification of relatively small numbers of two-locus (B-C and DRB1-DQB1) haplotypes suggested strong associations of those two loci, respectively. Five-locus haplotypes defined by high-resolution DNA typing correlated well with previously identified serology-based haplotypes in the population. The five most frequent haplotypes were: A*3303-Cw*1403-B*4403-DRB1*1302-DQB1*0604 (4.2%), A*3303-Cw*0701/6-B*4403-DRB1*0701-DQB1*0201/2 (3.0%), A*3303-Cw*0302-B*5801-DRB1*1302-DQB1*0609 (3.0%), A*2402-Cw*0702-B*0702-DRB1*0101-DQB1*0501 (2.9%), and A*3001-Cw*0602-B*1302-DRB1*0701-DQB1*0201/2 (2.7%). Several sets of allele level haplotypes that could not be discriminated by routine HLA-A, -B, and -DRB1 low-resolution typing originated from allelic diversity of A2, B61, DR4, and DR8 serologic groups. Information obtained in this study will be useful for medical and forensic applications as well as in anthropology.     

Analytical Evaluation of GeneXpert CT/NG,the First Genetic Point-of-Care Assay for Simultaneous Detection of Neisseria gonorrhoeae and Chlamydia trachomatis

GeneXpert CT/NG was evaluated with 372 characterized bacterial strains. Sensitivity of 10 genome copies/reaction was obtained for both agents. Four Neisseria mucosa and two Neisseria subflava isolates were positive for one of two gonococcal targets; however, the assay flagged all as negative. The assay was analytically highly sensitive and specific.     

HLA-A, -B及-DRB1等位基因的多态性与白血病易感性的关联

目的:探讨甘肃汉族人群中白血病的易感性与HLA基因多态性之间的关联。方法:采用序列特异性寡核苷酸探针杂交技术(PCR-SSO),对57例白血病患者和45例健康对照组进行HLA-A、-B及-DRB1基因分型。结果:在等位基因HLA-A、-B及-DRB1中,白血病患者的HLA-A01、-B38及-DR15基因的基因频率高于正常对照组(P<0.05),而HLA-A11及-DR03基因的基因频率明显下降(P<0.05)。结论:甘肃省汉族人群基因HLA-A01、-B38及-DR15对白血病具有遗传易感作用;而基因HLA-A11和-DR03对白血病具有遗传拮抗作用。     

Complete coding sequences and haplotypic associations of HLA-B*0707, -B*1524, -B*4405, -B*4802, -DRB1*0409, -DRB1*0411, -DRB1*1115, -DRB1*1305, and the novel allele -DRB1*0709. Group-specific amplification of cDNA from DRB1 alleles associated to DRB3 and DRB4

We present here the characterization of the complete coding sequences, previously unavailable, of the human leukocyte antigen (HLA) alleles B*0707, B*1524, B*4405, B*4802, DRB1*0409, DRB1*0411, DRB1*1115, DRB1*1305, and that of a new allele, DRB1*0709. For the isolation of cDNA from the DRB1 gene, we designed a novel set of polymerase chain reaction (PCR) primers that makes it possible to amplify separately the groups of DRB1 alleles associated to each of the DRB3 and DRB4 loci. The primary structures, functional features, evolutionary relationships, haplotypic associations, and population distributions of each of the nine HLA-B and -DRB1 alleles reported here are reviewed.     

Pelvic inflammatory disease isolates of Neisseria gonorrhoeae are distinguished by C1q-dependent virulence for newborn rats and by the sac-4 region.

The virulence mechanism of Neisseria gonorrhoeae in pelvic inflammatory disease (PID) is not well understood, and an objective diagnostic method to identify patients with PID is lacking. We investigated the hypothesis that development of PID was associated with a C1q-dependent virulence property of gonococcal strains. Recent development of a C1q-dependent experimental model of gonococcal infection (S. Nowicki, M. Martens, and B. Nowicki, Infect. Immun. 63:4790-4794, 1995) created an opportunity to evaluate this hypothesis in vivo. Therefore, the virulence of 32 clinical isolates (18 PID isolates and 14 local infection [LI] isolates) was evaluated in experimental rat pups. A serum bactericidal assay was used to characterize a gonococcal serum-resistant (ser(r)) phenotype. PCR primers designed to amplify a suitable-size gonococcal sac-4 DNA fragment (unique for serum-resistant donor JC1) were used to evaluate the association of serum-resistant genotype sac-4 with two phenotypes: C1q-dependent virulence expressed in vivo and resistance to bactericidal activity of human serum expressed in vitro. Strains were also characterized by auxotyping and serotyping. Of 32 gonococcal strains, 15 (46.7%) caused C1q-dependent bacteremia in rat pups and were sac-4 positive and ser(r). However, of the 15 isolates, 13 (87%) represented strains associated with human PID and 2 (13%) were associated with LI. None of the strains that were completely serum-sensitive (ser(s)) and sac-4 negative produced C1q-dependent bacteremia in rat pups, suggesting that both ser(r) and sac-4 were required for infection. The serum-resistant recombinant recipient of sac-4 produced C1q-dependent bacteremia in the rat model similarly to the serum-resistant donor of sac-4; the serum-sensitive parent strain did not produce bacteremia. These data suggest that sac-4-mediated serum resistance conferred C1q-dependent virulence and is a unique characteristic associated with PID. These newly identified features may contribute to the understanding of the pathogenic mechanism of PID-associated strains and open perspectives for establishing novel diagnostic methods.     

Gonorrhoea surveillance, laboratory diagnosis and antimicrobial susceptibility testing of Neisseria gonorrhoeae in 11 countries of the eastern part of the WHO European region

Quality-assured worldwide surveillance of antimicrobial resistance (AMR) in Neisseria gonorrhoeae is crucial for public health purposes. In the countries of the eastern part of the WHO European region the knowledge regarding gonococcal AMR is limited, and antimicrobials of many different types, sources and quality are used for gonorrhoea treatment. This study surveyed gonorrhoea incidence, laboratory diagnosis and gonococcal AMR testing in 11 independent countries of the former Soviet Union. The national gonorrhoea incidences remain mainly high. In general, gonococcal culture and AMR testing were rarely performed, poorly standardized and rarely quality assured. To establish a gonococcal AMR surveillance programme in Eastern Europe, i.e. the geographical area of the former Soviet Union, several actions have recently been undertaken by the Eastern European Sexual and Reproductive Health (EE SRH) Network and the WHO. The information provided herein will be useful in this respect.