Optimizing DNA Extraction Methods for Nanopore Sequencing of Neisseria gonorrhoeae Directly from Urine Samples

Empirical gonorrhea treatment at initial diagnosis reduces onward transmission. However, increasing resistance to multiple antibiotics may necessitate waiting for culture-based diagnostics to select an effective treatment. There is a need for same-day culture-free diagnostics that identify infection and detect antimicrobial resistance. We investigated if Nanopore sequencing can detect sufficient Neisseria gonorrhoeae DNA to reconstruct whole genomes directly from urine samples. We used N. gonorrhoeae-spiked urine samples and samples from gonorrhea infections to determine optimal DNA extraction methods that maximize the amount of N. gonorrhoeae DNA sequenced while minimizing contaminating host DNA. In simulated infections, the Qiagen UCP pathogen mini kit provided the highest ratio of N. gonorrhoeae to human DNA and the most consistent results. Depletion of human DNA with saponin increased N. gonorrhoeae yields in simulated infections but decreased yields in clinical samples. In 10 urine samples from men with symptomatic urethral gonorrhea, ≥92.8% coverage of an N. gonorrhoeae reference genome was achieved in all samples, with ≥93.8% coverage breath at ≥10-fold depth in 7 (70%) samples. In simulated infections, if ≥104 CFU/ml of N. gonorrhoeae was present, sequencing of the large majority of the genome was frequently achieved. N. gonorrhoeae could also be detected from urine in cobas PCR medium tubes and from urethral swabs and in the presence of simulated Chlamydia coinfection. Using Nanopore sequencing of urine samples from men with urethral gonorrhea, sufficient data can be obtained to reconstruct whole genomes in the majority of samples without the need for culture

TESTING PATTERNS FOR SYPHILIS AND OTHER SEXUALLY TRANSMITTED INFECTIONS IN PREGNANT WOMEN PRESENTING TO EMERGENCY DEPARTMENTS

Following an initial decrease in the incidence of congenital syphilis from 2008-2012, the rate of congenital syphilis rose by 38% across the United States between 2012-2014 (2). This trend followed a 22% rise in primary and secondary syphilis cases in women during the same period.(1) Vertical transmission of syphilis is a significant public health concern, contributing to stillbirth, infant mortality, and neurologic and skeletal morbidities in survivors. (2) The Centers for Disease Control and Prevention (CDC) recommends that all pregnant women be screened for sexually transmitted infections (STI) including HIV, syphilis, and hepatitis B at the first prenatal visit regardless of prior testing. The American College of Obstetricians and Gynecologists (ACOG) and the U.S. Preventive Services Task Force (USPSTF) also support similar recommendations. Yet, a CDC investigation into this epidemic revealed that 21% of women whose infants were diagnosed with congenital syphilis had no prenatal care, and of those who had at least one prenatal visit, 43% received no treatment for syphilis during pregnancy and 30% received inadequate treatment. (2, 3) Little is understood about factors associated with low STI screening during pregnancy in the US. In a 2014 study, Cha, et al. evaluated factors affecting the likelihood of STI screening in pregnant women in Guam. They found that the biggest barrier to STI testing was lack of prenatal care and insurance. Even women with access to prenatal care were not routinely screened for syphilis before 24 weeks’ gestation. Despite a 93.5% overall rate of screening for syphilis at any time during pregnancy, the authors found much lower screening 2 rates for other STIs, including 31% for HIV, 25.3% for chlamydia, and 25.7% for gonorrhea. (8) This suggests potential disparity in testing practices based on risk perception by providers or patients

Risk of pelvic inflammatory disease in relation to chlamydia and gonorrhea testing, repeat testing, and positivity: A population-based cohort study

Background: There is uncertainty around whether the risks of pelvic inflammatory disease (PID) differ following Chlamydia trachomatis (chlamydia) and Neisseria gonorrhoeae (gonorrhea) infection. We quantified the risk of PID associated with chlamydia and gonorrhea infection and subsequent repeat infections in a whole-population cohort. Methods: A cohort of 315123 Western Australian women, born during 1974–1995, was probabilistically linked to chlamydia and gonorrhea testing records and to hospitalizations and emergency department presentations for PID from 2002 to 2013. Time-updated survival analysis was used to investigate the association between chlamydia and gonorrhea testing, and positivity, and risk of PID. Results: Over 3199135 person-years, 120748 women had pathology test records for both chlamydia and gonorrhea, 10745 chlamydia only, and 653 gonorrhea only. Among those tested, 16778 (12.8%) had ≥1 positive chlamydia test, 3195 (2.6%) ≥1 positive gonorrhea test, and 1874 (1.6%) were positive for both. There were 4819 PID presentations (2222 hospitalizations, 2597 emergency presentations). Adjusting for age, Aboriginality, year of follow-up, health area, and socioeconomic status, compared to women negative for chlamydia and gonorrhea, the relative risk (adjusted incidence rate ratio) of PID was 4.29 (95% confidence interval [CI], 3.66–5.03) in women who were both chlamydia and gonorrhea positive; 4.54 (95% CI, 3.87–5.33) in those only gonorrhea positive; and 1.77 (95% CI, 1.61–1.94) in those only chlamydia positive. Conclusions: Gonorrhea infection conferred a substantially higher risk than chlamydia of hospitalization or emergency department presentation for PID. The emergence of gonorrhea antimicrobial resistance may have a serious impact on rates of PID and its associated reproductive health sequelae

Antimicrobial Resistance in Neisseria gonorrhoeae: Proceedings of the STAR Sexually Transmitted Infection-Clinical Trial Group Programmatic Meeting.

The goal of the Sexually Transmitted Infection Clinical Trial Group's Antimicrobial Resistance (AMR) in Neisseria gonorrhoeae (NG) meeting was to assemble experts from academia, government, nonprofit and industry to discuss the current state of research, gaps and challenges in research and technology and priorities and new directions to address the continued emergence of multidrug-resistant NG infections. Topics discussed at the meeting, which will be the focus of this article, include AMR NG global surveillance initiatives, the use of whole genome sequencing and bioinformatics to understand mutations associated with AMR, mechanisms of AMR, and novel antibiotics, vaccines and other methods to treat AMR NG. Key points highlighted during the meeting include: (i) US and International surveillance programs to understand AMR in NG; (ii) the US National Strategy for combating antimicrobial-resistant bacteria; (iii) surveillance needs, challenges, and novel technologies; (iv) plasmid-mediated and chromosomally mediated mechanisms of AMR in NG; (v) novel therapeutic (eg, sialic acid analogs, factor H [FH]/Fc fusion molecule, monoclonal antibodies, topoisomerase inhibitors, fluoroketolides, LpxC inhibitors) and preventative (eg, peptide mimic) strategies to combat infection. The way forward will require renewed political will, new funding initiatives, and collaborations across academic and commercial research and public health programs

Effect of Rectal Hygiene on Sexually Transmitted Infections Among HIV-Negative Men Who Have Sex with Men (MSM)

Abstract Background Rectal gonorrhea (NG) and chlamydia (Connecticut) infections are common among men who have sex with men (MSM). Rectal douching/enema (RDE) is a common practice among MSM that can affect the rectal microbiome. It is unclear if this practice is associated with acquiring rectal infections (RI) with either NG or CT. Methods From 2013–2015, 398 adult HIV-negative MSM and transwomen were enrolled in a randomized controlled study on text messaging for adherence to pre-exposure prophylaxis (PrEP). Participants were surveyed on sexual behavior, frequency of RDE, drug use, and nutritional habits in conjunction with routine sexually transmitted infection testing. Pearson’s χ 2 and two sample t-tests were used to measure significance of RDE and other risk factors with RI. Multivariable logistic regression model was used to control for confounding and assess the association of RDE with RIs. Confounders (i.e., age, number anal receptive sex, number sex partners) were selected a priori for inclusion in the final model based on a causal model and statistical significance. Results Of 397 participants, 262 (67%) performed RDE and 132 (33%) had at least one NG or CT rectal infection over 48 weeks. Number of condomless anal receptive sex acts (mean = 19, P < 0.001), condom use for anal receptive sex (P = 0.017), number of male sex partners in past 3 months (mean = 14, P = 0.001), and the use of poppers (P < 0.001) were associated with RI. There was no significant association between nutritional habits, probiotic foods or supplements and RI, with the exception of energy bars (P = 0.029). Controlling for confounders, RI was associated with RDE less than weekly with OR = 1.02 (95% CI 0.52–1.99) while RDE weekly or more had OR = 2.08 (95% CI 1.03–4.17). Stratified by number of partners, MSMs with more than the median (>6) number of partners had OR = 4.96 (95% CI 1.29–19.03) if performing RDE less than weekly, and OR = 6.03 (95% CI 1.55–23.49) if weekly or more. Conclusion Rectal hygiene with douching/enemas is a common practice among MSMs on PrEP, which increases the odds of acquiring rectal NG and/or CT. This finding is suggestive for the use of rectal hygiene products/practices as potential targets for sexually transmitted infection prevention. Disclosures All authors: No reported disclosures

Scoping Review On Gonorrhea Vaccines

Gonorrhea infection is caused by Neisseria gonorrhoeae and it is the second most common bacterial sexually transmitted infection around the globe. Although gonorrhea infection is treatable with antibiotics, the level of antimicrobial resistance continues to increase and has become a major public health concern. As options of available treatments are reduced by the emergence of AMR, it has become urgent to develop effective vaccines against gonorrhea infection. The purpose of this scoping review is to describe existing epidemiological evidence in support of gonorrhea vaccine development, to discuss the current vaccine candidates in clinical stages, and to evaluate the theoretical impact of gonorrhea vaccines. Published papers and clinical trial records on gonorrhea vaccine immunogenicity and efficacy, vaccine effectiveness and potential impact were reviewed after selection. Outer membrane vesicles-based meningococcal serogroup B vaccination showed moderate (approximate 30%) cross-protection against gonorrhea infections in multiple retrospective studies, and subsequent clinical studies assessing the immunogenicity and vaccine efficacy are currently undergoing. Modeling studies suggested that the theoretical impact of gonorrhea vaccines vary by vaccine efficacy, mode of action, duration of protection, uptake percentage, and disease prevalence in the population, but a vaccine with efficacy and duration of protection equivalent to the meningococcal serogroup B vaccine can have substantial impact and public health value if delivered in optimal vaccination strategy. However, generalizability of the study results remains questionable because studies included were mostly limited to developed country settings. To investigate vaccines with higher efficacy and to better evaluate the value of potential gonorrhea vaccines, research on disease natural history and gonorrhea epidemiology in low- or middle-income countries in the near future

Prevalence and determinants of Neisseria gonorrhoea and Chlamydia trachomatis infections in patients with pelvic inflammatory disease at Lusaka, Zambia

Background: Pelvic inflammatory disease (PID) is an inflammatory process affecting typically the endometrium, fallopian tubes, ovaries, parametrium and pelvic peritoneum with its surrounding structures. More than 85% of spontaneous PID is caused by sexually transmitted infections like gonorrhea, chlamydia, mycoplasma and others. WHO estimates that 340 million new cases of N. gonorrhea, Chlamydia and other sexually transmitted infections occur every year of which 85% are in developing countries. Gonorrhea and chlamydia account for 62 million and 92 million new infections respectively. In Zambia, 10% of outpatient attendance at health institutions is due to sexually transmitted infections. At University Teaching Hospital, pelvic inflammatory disease accounts for about 10 - 15% of gynaecological attendance. Despite these, there is no adequate information on magnitude and prevalence of these two organisms among patients with PID in Lusaka.Objective: The objective was to determine the prevalence and determinants of Neisseria gonorrhea and Chlamydia trachomatis infection among gynaecological patients diagnosed clinically to have PID at University Teaching Hospital, Lusaka. Several socio-demographic factors, sexual behaviour and clinical factors associated with chlamydia and gonorrhea were studied.Methodology: This was a descriptive cross sectional study conducted on women presenting clinically with pelvic inflammatory disease to the emergency outpatient gynaecological ward at UTH. Information about demographic, sexual behavioural history and clinical presentation was obtained using a structured questionnaire. Endo-cervical smear was obtained and screened for gonorrhea and chlamydia using respective immunoassay rapid test and gram stain for gonorrhea.Results: A total of 6 respondents were recruited out of which 43 (37%) had gonorrhoea while no chlamydia (0/6) was isolated. 4 (98.3%) had sexual partners while 2 had no sexual partner and no gonorrhea was isolated from them. Among those with sexual partners, 101(78.3%) had steady sexual partners, 19 (4.8%) had casual partners and 9 (7.0%) had anonymous sexual partners with 37 (36.6%), 10 (52.6%) and 6 (66.7%) gonorrhea isolation respectively. Some had multiple sexual partners. Gonorrhea was isolated from 4 (28.6%) of the 14 respondents who had one new sexual partner, and those who had two or more sexual partners had gonorrhea isolated. Gonorrhea isolation in relation to frequency of sexual intercourse per week was as follows: once 1/13 (7.7%), twice 2/11 (18.2%), thrice 3/11 (27.3%) and more than three times 32/65 (49.2%). Gonorrhea was high in those who had sex with casual or anonymous sexual partner under influence of alcohol 6/11 (54.5%) or obtained anonymous sexual partner from market, shopping centre 4/5 (80%), street, bar, disco, or night club 7/9 (77.8%). Regarding relationship to clinical features gonorrhea detection was as follows: 18/36 (50.0 %) for those with adnexial tenderness, 21/45 (46.7%) with inflamed cervix, 37/92 (40.2%) with lower abdominal tenderness and 32/87 (36.8%) with normally appearing cervix.Conclusion: The prevalence of Neisseria gonorrhea in gynaecological patients with PID at UTH in Lusaka was 37%. There was no chlamydia isolated. Low socioeconomic status and young age were the sociodemographic factors associated with a higher risk of gonorrhea. The sexual risk behaviours associated with gonorrhea were; the number of casual or anonymous sexual partners, frequency of sexual intercourse and non use of condoms. Lower abdominal pain and tenderness with cervical motion and adnexial tenderness were associated with higher gonorrhea isolation. There was no specific clinical predictor of either gonorrhea or chlamydi

An Investigation of the Effects of Alcohol Policies on Youth STDs

The purpose of this paper is to examine the role of alcohol policies in reducing the incidence of sexually transmitted diseases among youth. Previous research has shown that risky sexual practices (e.g., unprotected sex and multiple partners) that increase the risk of contracting a STD are highly correlated with alcohol use. If alcohol is a cause of risky sexual behavior, then policies that reduce the consumption of alcohol may also reduce the incidence of STDs. In this paper, we examine the relationship between alcohol policies (e.g., beer taxes and statutes pertaining to alcohol sales and drunk driving) and rates of gonorrhea and AIDS among teenagers and young adults. Results indicate that higher beer taxes are associated with lower rates of gonorrhea for males and are suggestive of lower AIDS rates. Strict drunk driving policies in the form of zero tolerance laws may also lower the gonorrhea rate among males under the legal drinking age.

Antibiotic-Resistant Gonorrhea: The Current Threat and Future Public Health Implications

Gonorrhea is a sexually transmitted bacterial infection with a high incidence rate in the United States and abroad. Untreated gonorrhea infections can lead to Pelvic Inflammatory Disease (PID), infertility in both men and women and blindness in babies infected by their mothers. The morbidity associated with gonorrhea, however, has been inhibited by the use of antibiotics, especially in the developed world. Since antibiotics were first used to cure gonorrhea in 1937, a variety of antibiotics have been used and subsequently discarded as first line drugs to treat the disease due to the gonorrhea bacterium’s adept ability to develop antibiotic resistance. Currently, the first line drug to treat gonorrhea is ceftriaxone, an injectable drug in the cephalosporin class of antibiotics. However, recent treatment failures after a standard dose of ceftriaxone, which have occurred in Japan, Australia and throughout Europe, have prompted experts’ concern about the future of gonorrhea treatment. While a variety of other antibiotics have been suggested as possible replacements for ceftriaxone, no known drug is a particularly good or immediate substitute, either because of insufficient research, a proven lack of efficacy or already-existing resistance within prevalent gonorrhea strains. In the absence of another antibiotic to take ceftriaxone’s place, the focus of public health professionals and clinicians must be on prevention. The Centers for Disease Control and Prevention (CDC), World Health Organization (WHO) and European Centre for Disease Prevention and Control (ECDC) should adjust their antibiotic-resistant gonorrhea working plans to prioritize prevention, rather than surveillance, in order to stem the tide of what could be a pandemic of an untreatable bacterial infection, the likes of which have not been experienced in the modern antibiotic era. The public health consequences of such an epidemic would be severe with global increases in morbidity, costs and Disability Adjusted Life Years (DALYs) lost due to gonorrhea infection