Chlamydia: The Female Reproductive System and Infertility

Abstract

Chlamydial infection can cause diseases in many organs, including the genitourinary system. It is the most reported sexually transmitted bacterial infection throughout the world and one of the leading cause of female infertility. Chlamydia affects columnar epithelium, so adolescent women are particularly at risk since the squamocolumnar junction is located on the ectocervix until early adulthood. The bacterium is usually transmitted through sexual activity. Genital tract infection is the most common clinical picture but 50% of infected men and 80% of infected women are asymptomatic. This is the most important reason for the infection’s being unrecognized and untreated. The most significant morbidity related to infection is partial or total sterility due to obstruction and scarring of the fallopian tubes. Chlamydia trachomatis infection, even if it does not present clinical symptoms, has been shown to be associated with increased tubal factor infertility, implantation failure, and disruption of embryo development.

1. Introduction

Chlamydias are small gram-negative, obligate intracellular living microorganisms, preferably infecting squamocolumnar epithelial cells. The microorganisms can be divided into two subtypes as Chlamydia (e.g., C. trachomatis) and Chlamydophila (e.g., Chlamydophila pneumoniae and Chlamydophila psittaci). C. trachomatis is divided into 19 different serological variants (A, B/Ba, C, D/Da, E, F, G/Ga, H, I/Ia, J, K, L1, L2, L2a, and L3) according to monoclonal antibody-based analyses and variants that are classified according to ompA genotyping. Of these, types A, B, Ba, and C are the causative agents of Trachoma, an endemic serious eye disease in Africa and Asia; D-C strains lead to genital tract infections. L1-L3 strains cause Lymphogranuloma venereum disease, which is especially seen in tropical countries and is characterized by genital ulcers [1].

Chlamydia can affect various organs, and the genitourinary system is one of the major sites of this infection. It can cause clinical conditions in nasopharynx, epididymis, urethra, cervix, uterus, and salpinx [2, 3, 4]. It is the most reported sexually transmitted bacterial infection and one of the major causes of female infertility. This infection also leads to conjunctivitis, pneumonia, afebrile pneumonia syndrome (in vaginally born babies from infected mothers), and trachoma, a leading cause of acquired blindness in the world and perihepatitis condition also known as Fitz-Hugh-Curtis syndrome [5].

There are one million sexually transmitted disease transmissions every day in the world [6]. The annual number of C. trachomatis genital infections reached 4 million annually in 2018, up from about 2.86 million cases in 2008 [7]. Research shows similar incidences in Germany [8], France [9], the Netherlands [10], New Zealand [11], and Australia [12]. In a report by the World Health Organization (WHO) Initiative for Vaccine Research (IVR) it is estimated that there are more than 140 million cases of C. trachomatis infection worldwide [13]. Chlamydia carrier rates in the sexually active female population are around 20%. This ratio is 2–3 times higher than the N. gonorrhea incidence.

2. Pathophysiology

Chlamydia affects columnar epithelium, so adolescent women are particularly at risk since the squamocolumnar junction is located on the ectocervix until early adulthood. Chlamydia has extraordinary features regarding its life cycle. It has two different forms in the course of infection in both intracellular and extracellular environments. The elementary body (EB) is the infectious form existing in the extracellular space; it is a spore-like, inactive structure that enters the host cell. Inside the cell, it turns into an active form called a reticulate body (RB). The RB uses the host cell’s amino acids and the energy sources in the form of ATP to synthesize its own DNA, RNA, and proteins to replicate and after enough RBs have formed, some of them turn back to the EB form, which can then exit the initial host cell and infect others. This cycle is then repeated in the adjacent cells [14, 15]. Thus, this process creates an immunogenic environment around the infection. However, there are ways for C. trachomatis to escape and evade the immune system. For example, by preventing T-cell immune recognition, it down-regulates the major histocompatibility complexes I and II; it modulates some specific cytokines, such as beta interferon, type 1 interferons, interleukin 18 and, inhibits apoptosis by the secretion of Chlamydial protease-like activity factor proteins and enhancing cell survival signals. By this way, it creates a chronic inflammation that allows the infection to become persistent. Replication mechanisms that adapt to the environment during the biphasic development cycle and evolutionary defense mechanisms that allow to escape from the immune system and environmental inflammatory stress are also challenging obstacles to infection treatment and vaccine development [16]. Still, with advances in silico studies using bioinformatics tools and machine learning-based modeling, Shiragannavar and his colleagues, have succeeded in developing a candidate vaccine that stimulates T and B cells in a way that provides long-term immunity [17]. There are also other vaccine candidates that use immune and proteomic approaches as well as in silico methods [18].

The bacterium is usually transmitted through sexual activity. The risk of an infected man infecting an uninfected woman with each sexual contact is 25%. Chlamydia can also spread vertically. The risk of transmission from the infected mother to the newborn is around 50–60%, and in most cases, the neonatal infection is in the form of conjunctivitis or pneumonia. (In 10–20% of cases, in the form of Afebrile Pneumonia Syndrome).

Genital tract infection is the most common clinical picture. The incubation period is around 1–3 weeks. In total, 50% of infected men and 80% of infected women are asymptomatic. But the infection can cause mucopurulent cervicitis in women, and it can lead to urethritis in men [19]. Ascending infection can result in the development of PID in women and is the most common cause of epididymitis in men under 35 years of age. In total, 5–10% of women who have had PID may progress to perihepatitis also known as Fitz-Hugh-Curtis syndrome.

Although the presence of any STD in one patient increases the likelihood of coinfection with another STD, the most common coinfection in such a case is the combination of Chlamydia and Gonorrhea. In total, 40% of women and 20% of men with chlamydia infection are coinfected with gonorrhea [20, 21]. Other pathogens that can cause coinfection with Chlamydia are Mycoplasma genitalium [22, 23] and HPV [24, 25] possibly causing an association between cervical intraepithelial neoplasia and Chlamydia [26]. The frequency of Reiter’s syndrome (reactive arthritis, conjunctivitis, and urethritis) in patients with chlamydia is also increased compared to the normal population. LGV cases are the reason for 10% of genital ulcers in tropical countries. In the course of a LGV case, localized inguinal lymphadenopathy and ulceration develop within 2–12 hf after exposure to infection. In untreated cases, proctitis, rectal strictures, and elephantiasis secondary to lymphatic obstruction may occur.

3. Etiology

C. trachomatis transmission occurs with direct contact, this includes the vaginal, anal, or oral sexual routes and vertical transmission to the newborn from an infected mother during vaginal birth.

Specific risk factors for the chlamydial infection are 15–24 years of age, poor socioeconomic conditions, multiple sexual partners, exchange of sex for money, intercourse without a barrier contraceptive, current coinfection with another STD, or a history of previous STD, certain cytokine polymorphisms that are related to severe disease and the risk of tubal factor infertility [27], specific variants in Toll-like receptor 1 and 4 genes creating infection predisposition [28].

4. Prognosis

Antibiotic therapy is 95% effective as a first-line treatment. If treatment is started early and fully completed, the prognosis is good. Although treatment failure with first-line treatments is quite rare, recurrences may occur with alternative treatments. Reinfection is very frequent, either because the partner is not treated or because it is reacquired from a new partner. Therefore, treatment of all possible sexual partners is mandatory.

Abscess rupture due to salpingitis and progression to tubo-ovarian abscess and death due to peritonitis are rare. Chlamydia is an indirect cause of ectopic pregnancy-related deaths. Death due to ectopic pregnancy caused by chlamydia is more likely than deaths due to tubo-ovarian abscess.

5. Clinic considerations

In Chlamydia, due to the asymptomatic course of the infection, diagnosis can usually be delayed until discovery of a symptomatic partner or a positive screening result. Therefore, the chlamydia screening programs, which have been shown to reduce PID rates [29, 30], can be claimed to be necessary for the timely diagnosis and treatment of this infection.

Despite this mostly asymptomatic course, C.trachomatis can cause a broad range of urogenital diseases, including cervicitis, endometritis, salpingitis, pelvic inflammatory disease, urethritis, prostatitis, epididymitis, lymphogranuloma venereum; and extragenital diseases like conjunctivitis, pharyngitis, reactive arthritis, proctitis, and neonatal pneumonia. Asymptomatic men and women act as reservoirs for the infection. In a previous study, the rate of transmission was estimated to be 68% both for women and men [31].

In the symptomatic patients, clinical signs and symptoms depend on the infection site, and local mucosal inflammation with a subsequent discharge can lead to vaginitis, cervicitis, and urethritis in females and urethritis in males.

Symptoms are felt with different severity in different anatomic regions, depending on the bacterial variants that vary according to the specific epitopes encoded by ompA [32, 33]. There is a link between the genotype of C.trachomatis and its pathogenicity and the severity of infection [34, 35]. Chen et al. showed in their study that patients infected with genotype D, the most prevalent type in their study, had a lower risk of both coinfection with other pathogens and cervical cancer. Genotype F was found to be the most associated with bacterial coinfections, and serovar G was also the type at risk for coinfection. Genotype E was associated with mucopurulent cervicitis and cervical dysplasia [36]. Extensive studies of C. trachomatis serovars have found that variant prevalence show marked geographic distributions and differ by studied region, gender, ethnicity, and sexual orientation [34, 37, 38, 39, 40].

The most common signs, symptoms, and history issues can be listed as follows:

These can be encountered in both sexes:

• Possible history of STDs.

• Dysuria.

• Urethral mucopurulent discharge.

These can be seen in females:

• History of sexual activity without a barrier contraceptive method or with the failure of the method.

• Vaginal discharge.

• Dyspareunia.

• Dysuria.

• Abnormal vaginal bleeding (postcoital or intermenstrual).

• In case of receptive anal intercourse; rectal discharge, proctitis, or both.

• Lower abdominal pain.

• Fever (in PID).

• No symptoms (in 80%).

Signs of chlamydial infection in women may include the following:

• Mucopurulent cervical or vaginal discharge.

• Cervical friability (easy bleeding on manipulation).

• Cervical motion tenderness.

• Urethral discharge (usually thin and mucoid).

• Mucopurulent rectal discharge.

• Adnexal or lower abdomen fullness or tenderness (progression to PID).

• Upper right quadrant abdominal tenderness (Fitz-Hugh-Curtis syndrome).

Signs of lymphogranuloma venereum (LGV) may include the following:

• Genital ulceration.

• Localized inguinal adenopathy or buboes.

• “Groove sign” – Separation of the inguinal and femoral lymph nodes by the inguinal ligament (seen in 15–20% of patients).

6. Complications

Chlamydial infections are one of the most important causes of female infertility. Since infertility is one of the most significant issues to focus on chlamydia, it will be discussed in a separate section.

C.trachomatis is also the leading cause of PID. PID is a serious condition that may require hospitalization, intravenous antibiotic therapy, and tests to rule out a tubo-ovarian abscess. The risk of ectopic pregnancy is 7–10 times higher in women who have had PID than in those who have not. Pelvic adhesions involving the ovaries and tubes in 15% of women after PID may cause chronic pelvic pain in the long term. The case of perihepatitis, also known as Fitz-Hugh-Curtis syndrome, is a rare complication of PID, which is five times more common in chlamydia than in N. gonorrhea. Especially with serotype G infection, the risk of developing cervical cancer increases by about 6.5 times. Chlamydia infection also increases genital mucosal inflammation, facilitating HIV transmission.

A pregnant woman with a chlamydia infection can pass the infection to the baby during childbirth, and this can lead to pneumonia or conjunctivitis in the baby. Neonatal conjunctivitis, if remains untreated, can lead to blindness. Reiter’s syndrome, reactive arthritis that may develop secondary to the immune response after primary chlamydia infection, is manifested by asymmetrical polyarthritis, urethritis, uveitis, mouth ulcers, circinate balanitis, and keratoderma blennorrhagica. The etiology is uncertain, but it usually follows an infectious attack and 80% of patients are HLA B27 positive. Other serious potential complications that can be related to chlamydia infection are miscarriage [41] and preterm birth [42].

There is no known association between chlamydia and tumor development in men, according to the evidence to date. However, this relationship in women has been reported by some authors. Paavonen et al. showed that the presence of enhanced antibodies to heat shock protein-60 is a high risk for cervical cancer; this also suggests that persistent C. trachomatis infection is associated with the development of cervical tumors [43]. One of the possible molecular mechanisms explaining the association of chlamydial infection with increased cervical cancer risk is that the infection generates an inflammatory response that triggers releases of ROS, cytokines, chemokines, growth, and angiogenic factors, leading to genetic instability and abnormal mitosis [44, 45]. C. trachomatis also affects beta-catenin and N-cadherin proteins, which have significant structural and regulatory roles [46]. In addition, there is evidence that CT infection increases HPV transmission and persistence. This coinfection increases the risk of cervical cancer as the epithelial destruction caused by the bacterium facilitates the entry of the virus; at the same time, with the weakening of the immune system, a microenvironment is formed that prepares the ground for the development of cancer [47, 48]. Anttila et al. found that the type of bacteria which bears the highest risk of developing cancer is the G serotype [49].

Since persistent inflammation is known to be related with tumor development, the ovaries may also be affected by chlamydia infection, as expected [50]. Shanmughapriya and colleagues showed that about 80% of ovarian cancer patients are infected with chlamydia [51]. Correspondingly, several studies have shown that the anti-Hsp60 protein is increased in ovarian cancer patients [52]. But despite these evidence, there are different consequences related to this issue, so more studies are needed to shed light on this topic [53].

7. Differential diagnosis and diagnostic workup

Since Chlamydia trachomatis can cause a wide spectrum of clinical presentations and manifestations, differential diagnosis of various signs and symptoms are also broad. Similar clinical manifestations affecting each body part should be considered carefully for other etiologies. The most common diseases that should be considered in differential diagnosis can be listed as: vaginal candidiasis, bacterial vaginosis, gonorrhea,Trichomonas vaginalis, Ureaplasma or Mycoplasma genitalium infection, foreign body, genital herpes, urinary tract infection, appendicitis, constipation, ovarian cysts, endometriosis adenomyosis, inflammatory bowel disease, allergy, cervical or endometrial polyp, cervical cancer, cervical ectropion, syphilis, chancroid, granuloma inguinale, leiomyoma, and pregnancy.

Considering the C. trachomatis infections, trachoma is the only one that can be diagnosed based upon clinical findings. All other chlamydial infections require laboratory confirmation. All patients with any sexually transmitted disease (STD) should also be evaluated for chlamydial infection because of the possibility of coinfection. Based on the patient’s sexual practices achieved from history; endocervical, urethral, rectal, or oropharyngeal specimens should be collected and evaluated for C. trachomatis infection in both males and females [54]. Today, the gold standard for the diagnosis of urogenital chlamydia infections is nucleic acid amplification testing (NAAT) [55, 56]. A voided urine sample, whether first-void or midstream can be used to detect the chlamydial organism for nucleic acid amplification testing (NAAT). Self-collected vaginal swab specimens are shown to be equivalent in sensitivity and specificity to those collected by a clinician using NAATs [55, 57, 58]. If the newly mother had a documented untreated chlamydial infection during pregnancy, the infant should be treated doubtlessly without a need for confirmation.

7.1 Basic laboratory tests

A complete blood count (CBC) must be done if pelvic inflammatory disease (PID) is suspected. HIV testing, testing sexual partners for Chlamydia and a Pap smear test should also be considered. A pregnancy test is essential for females when determining the treatment because pregnancy is a contraindication for some treatment options.

7.2 Cytology and cell culture

Cytologic diagnosis is used to evaluate endocervical scrapings in genitourinary infection, but cultures are difficult to gather, not easy to analyze, and their sensitivity is low, many false-negative results are encountered. Because of the need for expert laboratory skills, they are also expensive. They may be incompatible when trying to assess a large number of patients. But they are still mandatory in certain clinical situations such as legal indications like rape or sexual abuse because of their high specificity (100%). Cell culture can also be the choice for rectal specimens because of the confounding effect of the stool microorganisms in other tests when interpreting the results.

C. trachomatis can be grown well in cell lines like McCoy and HeLa cells. Incubation time is 40–72 hours, depending on the cell type and specific biovar. Intracytoplasmic inclusions can be captured either by Giemsa stains or by immunofluorescent staining with monoclonal antibodies.

7.3 Molecular techniques for detecting antigen, DNA, or RNA/rapid tests

Since C.trachomatis only grows within columnar cells, it is essential to gather a specimen directly from the cervix or urethra that will involve cells. When trying to obtain cells from vaginal or urethral discharge, it should be tried to apply pressure to the inside of the cervix or urethra. In males, after the urethra is milked down for secretions, collection swabs should be inserted 1–2 cm inside to urethra or “kissing slide” method can be used for sample collection [59]. Transport and kit’s manufacturer instructions should always be fulfilled.

Enzyme-linked immunosorbent assay (ELISA) is the most preferred test for Chlamydia in outpatient clinics and emergency departments for large number of patients since it is cost-effective and mostly automatized. It has 40–60% sensitivity and a 99% specificity.

Direct fluorescent antibody (DFA) testing for C. trachomatis has a sensitivity of 50–80% and a specificity of 99%. It is often preferred to confirm other assays but, labor and skilled personnel are needed to perform.

7.4 Nucleic acid amplification tests

Non-culture tests for the detection of C. trachomatis have been substantially replaced by higher-performing NAATs. These tests show high performance even in noninvasive samples. NAATs have recently become the test of choice to effectively screen and diagnose infection because of their high sensitivity and their ability to perform noninvasive testing that does not require pelvic examination or urethral swab [60, 61, 62]. NAATs target and amplify nucleic acid sequences found in almost every clinical strain of C. trachomatis, including genital, LGV, and ocular serovars. The APTIMA Combo 2 Assay used for ribosomal RNA can be used on liquid-based Pap smear samples [63]. The gold-standard method for bacterium genotyping is DNA sequencing of the ompA gene, encoding the major outer membrane protein (MOMP) [64]. This method is relatively simple and inexpensive. Most studies have reported sensitivity of more than 70% and specificity of 97–99% in populations where the prevalence of infection in men and women is 5% or more. The FDA-approved NAATs are recommended for the detection of infections caused by C.trachomatis and N.gonorrhoeae in men and women with or without symptoms [56]. Multiplex polymerase chain reaction assays are now started to be used widely, especially for polymicrobial infections and simultaneously testing the possible STDs from a single specimen [65, 66]. Older non-culture or non-NAAT tests with lower sensitivity are not recommended anymore. The Centers for Disease Control and Prevention (CDC) recommend NAATs for extragenital sites such as rectal and oropharyngeal infections because of their higher sensitivity and ease of sample handling and processing. Routine repeat testing is not recommended for NAAT-positive genital tract infections because repeated testing does not increase the positive predictive value of the test. Cultures of C. trachomatis and N. gonorrhoeae may still be necessary for the detection of sexual abuse in boys and extragenital infections in girls [56].

While NAATs are sensitive, they also have some drawbacks. Primarily, they are expensive, and so healthcare units may not be able to use them for comprehensive screening due to cost [67]. Another disadvantage of FDA-approved NAATs is that they cannot distinguish LGV strains from others. This is an important point because the duration of treatment in LGV infections needs to be extended. In addition, NAATs detect the DNA or RNA of the bacterium rather than the live microorganism, and it is common for control tests 3 weeks after the end of treatment to still show positivity [68]. For this reason, NAATs should not be used as a cure-determination test except in pregnant women, who must be shown to have a cure 3–4 weeks after the end of treatment to prevent infection in the infant.

In studies to shorten test result times, a prototype developed by TwistDx (prototype TwistDx RPA assay, Cambridge, UK) with an isothermal recombinant polymerase amplification approach shows promise. CT detection with this method takes 15 minutes. Validation studies of this prototype are still ongoing. But if this test is approved and made commercially available, it will mark a milestone in CT infection control, allowing doctors to diagnose and treat it in the same session [69, 70].

7.5 Serology

Antichlamydia immunoglobulin M (IgM) positivity is not common in adults with genital tract infections. Antichlamydial immunoglobulin G (IgG) positivity is high in sexually active adults, even if there is no active infection, and this most likely indicates a previous infection. Although there is a statistically significant relationship between chlamydia-specific serum immunoglobulin A (IgA) and active infection, none of the serological tests are sufficient to detect active disease in terms of clinical sensitivity, specificity, and predictive values. Therefore, it is not recommended for the diagnosis of genital tract infections. Nevertheless, they are still being used for research purposes and appear to be useful, especially for the detection of past infections using IgG.

7.6 CT, radiography, and ultrasonography

Imaging techniques are usually not necessary for uncomplicated genital chlamydia infections. However, CT and ultrasonography may be useful in complicated upper genital tract infections. For example, Fitz-Hugh-Curtis syndrome (perihepatitis) can be diagnosed with CT, and ultrasound can be used to investigate the presence of a tubo-ovarian abscess.

8. Screening

The social and financial burdens imposed by chlamydia are quite numerous. For this reason, some states such as the UK, Australia, the Netherlands, and Sweden have established a national chlamydia screening program. Each of these programs aimed to reduce the transmission of infection and its overall prevalence in the community with different strategies [71, 72]. Such a program should cover all sexually active individuals who are at an age where clinical intervention can alter long-term outcomes [73]. Evidence supports that screening should be available to those under 25 years old [74]. Studies on the subject have shown that because it is less invasive and easier, the postal screening method, in which people take samples on their own instead of the traditional way in the medical setting, increases screening rates [75, 76]. As Hoenderboom et al. points out, these samples may not be blood, but urine or vaginal swab samples [77].

Despite different opinions on the cost-effectiveness of screening programs, some authors argue that the screening program in the UK should be supported in this respect as well [78, 79, 80]. Retrospective studies of the chlamydia screening program in Sweden have shown that even in the first phase, the number of new cases has decreased. However, this reduction was achieved by using a more precise testing method, such as PCR, instead of more traditional methods such as culture. It was therefore concluded that faster, easier and more sensitive methods should be used for the diagnosis of chlamydia [37, 80]. Some other studies reported that the number of chlamydia infections was higher than expected, so screening should be done regardless of the estimated prevalence [81, 82]. In addition, several studies report that the prevalence of infection is high in adolescents and young women due to both their biological and behavioral predispositions, so a screening program should be established for at least 15–24 years of age [83]. All this evidence shows that it is important to establish screening programs to cover all sexually active individuals and to repeat tests for possible recurrent infections at regular intervals in order to prevent the spread of chlamydia and possible morbidity in the community by treating infected people in a timely manner. For such a screening program to be successful, noninvasive screening methods must be used. In this way, more people will be reached and the real incidence in the population will be determined, the measures to be taken for the control of the infection will be planned in a healthier way and the society will be made aware of the risk factors through prevention campaigns [84].

Recently, Huai and colleagues published a meta-analysis to estimate the prevalence of chlamydia infection worldwide and found that rates varied widely in the regions they studied, with the lowest prevalence in Southeast Asia. The authors then concluded, based on previous studies on the cost-effectiveness of screening programs, that it is critical to establish Chlamydia trachomatis screening programs using different guidelines in Latin America and Africa [85].

The US Preventive Services Task Force also recommends routine Chlamydia screening for sexually active young women to prevent consequences of undiagnosed and untreated chlamydial infection and has made the following recommendations: [86, 87].

• Screen for chlamydial infection in all sexually active nonpregnant young women aged 24 years or younger and for older nonpregnant women who are at increased risk.

• Screen for chlamydial infection in all pregnant women aged 24 years or younger and in older pregnant women who are at increased risk.

• Do not routinely screen for chlamydial infection in women aged 25 years or older, regardless of whether they are pregnant, or if they are not at increased risk.

9. Treatment

The most important points in the treatment of chlamydia infection are to make the correct diagnosis and to ensure the patient’s compliance with the treatment. Undiagnosed chlamydia infection can progress to PID and result in partial or complete infertility. It is undesirable for this to happen early in life, before childbearing. STIs can often be confused with a urinary tract infection. Therefore, those with a history of recurrent urinary tract infections should be evaluated for STDs. Adolescents are a high-risk group for noncompliance with treatment, especially if they are trying to keep secret from their parents. In this group, single-dose, in-office treatment is increasingly used to ensure compliance and confidentiality. Partner treatment is vital for the prevention of reinfection. The development of PID in an adolescent should be considered as an absolute indication for hospitalization due to noncompliance with prolonged treatment regimens and the possibility of developing infertility. Before starting treatment, samples should be taken from the infection area for laboratory or culture examination, and a pregnancy test should be performed as it may change the treatment and follow-up plan. Antibiotic therapy should be started as soon as possible. Compliance with treatment, cost and potential side effects should be considered in the selection of treatment, and the possibility of coinfection of gonorrhea should be kept in mind. It should be reminded that sexual intercourse should be avoided until treatment is complete and all sexual partners have been tested for infection.

Treatment should be started as soon as genitourinary chlamydial infection is diagnosed or suspected. Chlamydias are sensitive to antibiotics that affect DNA and protein synthesis; these include tetracyclines, macrolides, and quinolones [88]. The CDC recommends azithromycin and doxycycline as first-line drugs for the treatment of chlamydia [55, 60]. Medical treatment with these agents is 95% effective. Alternative medicines include erythromycin, levofloxacin, and ofloxacin [55]. Since the FDA issued a warning in 2013 that azithromycin can cause life-threatening arrhythmias, doxycycline should be preferred in patients with QT-interval anomalies or taking antiarrhythmic drugs. There is no need to retest for a cure after treatment, but a reevaluation is recommended after 3 months due to the high probability of reinfection [60].

In cases where compliance to treatment may decrease due to reasons such as cost, age, and confidentiality, it is recommended to apply single-dose treatment under observation for lower genital infections.

Due to the severity of potential complications, the presence of a condition affecting the upper genital tract should be carefully and meticulously investigated, especially in adolescents. Improper and inadequately treated PID can result in chronic pelvic pain, infertility, and sepsis. Monitoring of hospital treatment and response to treatment is especially important when PID is suspected, as adolescents may have trouble ignoring symptoms and continuing follow-ups.

In the treatment of PID, even if it is known gonorrhea to be present, treatment against C.trachomatis and anaerobic bacteria should always be included. Oral and parenteral regimens have shown similar efficacy in mild or moderate PID [60]. Patients treated in the hospital should not be discharged until significant clinical improvement is seen and confirmation that the patient will complete medical treatment. The recommended parenteral regimens are a continuation of doxycycline with cefoxitin or cefotetan for 14 days. Alternatively, clindamycin-gentamicin or doxycycline plus ampicillin-sulbactam may be given.

Out-of-hospital treatment for PID; following a single dose intramuscular administration of a second or third-generation cephalosporin, administration of doxycycline for 14 days, with or without metronidazole 500 mg for 14 days. After the emergence of quinolone-resistant cases of N. gonorrhoeae, treatments containing quinolone are no longer recommended for the treatment of PID. For sexual partners of the index case, treatment should also be given if the last sexual intercourse was within the last 60 days. Patients undergoing treatment for gonorrhea should also be treated for chlamydial infection.

Regarding treatment during pregnancy, CDC guidelines recommend a single dose, 1 g of azithromycin. Alternatively, amoxicillin 500 mg or erythromycin three times a day for 7 days can be used. Doxycycline, ofloxacin, and levofloxacin are contraindicated in pregnancy. To demonstrate eradication of chlamydia in pregnancy, it is recommended to test 3–4 weeks after the end of treatment, preferably by the NAAT method.

9.1 Treatment failure and novel approaches

The main causes of treatment failure are noncompliance with treatment, early testing for cure, and reinfection as a result of sexual partners not being adequately informed and adequately treated. In addition, treatment failure can be caused by antibiotic resistance caused by gene mutations in the bacterium or by the bacterium becoming insufficiently cleaned and persistent due to its natural characteristics [89]. In an in vitro study of antibiotic resistance, which included Croatia, the country with the highest azithromycin consumption in Europe, resistance was not shown to either azithromycin or doxycycline [90]. However, an experimental study in the UK comparing azithromycin with doxycycline found that treatment failure with azithromycin was higher in nongenital infections [91]. Multidrug-resistant CT serovars may be the reason for the ineffectiveness of azithromycin therapy. Some in vitro studies show that point mutations in the ribosomal proteins of the bacterium L genotype are responsible for azithromycin resistance [92]. There is also in vitro evidence that previous penicillin exposure may lead to azithromycin resistance in C. trachomatis [93].

Based on this evidence, it is clear that new drugs are needed to be developed to successfully combat C. trachomatis infection. Some researchers have studied Corallopyronin A, an antimicrobial compound synthesized by Corallococcus coralloides, and have shown that it inhibits CT proliferation [94] Shima et al. have also found promising results with this compound and have proposed it as an alternative to CT treatment in the future [94, 95]. A nanoparticle developed by Yang et al. successfully prevented vaginal CT infection by stimulating autophagy in human cells [96]. Recently, Nunez-Otero and his team demonstrated the role of a second-generation 2-pyridone amide molecule (KSK213) in the control of CT infection, in which they reduced its toxicity without damaging the commensal flora. This molecule acts through transcription inhibition in critical genes responsible for the conversion of EB to RB, the key point in the CT infection cycle [97].

In addition to all these, natural anti-chlamydia treatments derived from herbal extracts are also emphasized. Hamarsheh et al. studied the effect of Artemisia Inculta Delile extract and showed that it effectively inhibits infection in Hela cells [98]. As the issue of antibiotic resistance remains critical since 2020, some researchers have studied potential nonantibiotic agents. Lam et al. published their findings on cyclic peptomers that inhibit gram-negative bacteria and recommended 4EpDN cyclic peptomer as a prophylactic treatment against chlamydia [99].

Drug repurposing, which has been researched mostly in cancer treatment, has also been tried in this regard. Itoh et al. have shown that Bortezomib, an anticancer drug, may also be effective in treating CT infection through the induction of apoptosis [100]. More comprehensive studies are needed to be able to apply all these new strategies in the treatment of CT infection and to put the results of research into clinical practice. In order to eradicate this infection worldwide, the development of an effective vaccine in addition to all treatment strategies is critical.

9.2 Posttherapy care

Due to the high incidence of reinfection, retesting is recommended in the third month after treatment of chlamydia, gonorrhea, and trichomonas. In pregnancy, control testing after amoxicillin and erythromycin treatment should be considered. Due to the positive results from nonviable organisms, it would be better not to use non-culture methods in control tests.

Patients should be reminded to refrain from sexual activity for 7 days after a single dose treatment, and for longer treatments until the end of treatment and all sexual partners have been treated.

10. Prevention

Sexually active people should be aware of all other STDs, not just genital chlamydia infection. Diagnosed patients should be checked for all other STDs as much as possible. If possible, all sexual partners should be referred for diagnosis and treatment. Patients should also be informed that the most important way to prevent these infections, other than avoiding sexual activity, is to practice safe sex, that is, to use an appropriate barrier method such as a latex condom in every sexual relationship.

The American College of Obstetricians and Gynecologists (ACOG) has released guidelines about expedited partner therapy for chlamydial and gonorrheal infections [101, 102]. Although developed to prevent reinfection of chlamydia and gonorrhea, the recommendations may also be used for other STDs.

The ACOG recommendations can be listed as follows:

• Expedited partner therapy to prevent reinfection, with the legalization of expedited partner therapy.

• Counsel partners to undergo screening for HIV infection and other STDs.

• Expedited partner therapy is contraindicated in cases of suspected abuse or compromised patient safety, pretreatment evaluation for abuse potential is recommended.

• Expedited partner therapy medications and protocols based on CDC, state, and/or local guidelines.

11. WHO guidelines on the treatment of Chlamydia trachomatis infection

11.1 Uncomplicated genital chlamydia

WHO recommendations for the treatment of uncomplicated genital chlamydia are as follows [103]:

• Azithromycin 1 g orally as a single dose or.

• Doxycycline 100 mg orally twice a day for 7 days or one of these alternatives: tetracycline 500 mg orally four times a day for 7 days, erythromycin 500 mg orally twice a day for 7 days, or ofloxacin 200–400 mg orally twice a day for 7 days.

11.2 Anorectal chlamydial infection

In anorectal chlamydial infection, the WHO recommends doxycycline 100 mg orally twice a day for 7 days over azithromycin 1 g orally as a single dose.

11.3 Chlamydial infection in pregnant women

WHO recommendations for the treatment of chlamydial infection in pregnancy are as follows:

• Azithromycin recommended over erythromycin,

• Azithromycin recommended over amoxicillin,

• Amoxicillin recommended over erythromycin,

• Azithromycin 1 g orally as a single dose or,

• Amoxicillin 500 mg orally three times a day for 7 days or,

• Erythromycin 500 mg orally twice a day for 7 days.

11.4 Lymphogranuloma Venereum

WHO recommendations for the treatment of lymphogranuloma venereum (LGV) are as follows:

• In adults and adolescents with LGV, the guidelines suggest doxycycline 100 mg orally twice daily for 21 days over azithromycin 1 g orally weekly for 3 weeks.

• Good practice dictates the treatment of LGV, particularly for men who have sex with men and for people with HIV infection.

• When doxycycline is contraindicated, azithromycin should be provided.

• When neither treatment is available, erythromycin 500 mg orally four times a day for 21 days is an alternative.

• Doxycycline should not be used in pregnant women.

12. Infertility: a major complication of C. trachomatis infection

Potential factors affecting the association between chlamydia infection and infertility in women can be summarized in four categories; [1].

1. Host factors: Behavior, prevalence, genotype, microbiome.

2. Immunological factors: Cellular pathology, cHSP60 antibodies, suppressed immunity, IFN-gamma production.

3. Epidemiological factors: Age, sexual behavior, smoking, recurrent infection.

4. Pathogenic factors: Serovar, infectious burden, persistence, genotype, treatment failure, ability to ascend.

As stated in a review examining the sexually transmitted disease and infertility association, tubal factor infertility (TFI) is one of the most common causes of infertility. While it is responsible for up to 33% of female infertility cases worldwide, this rate is disproportionately high in developing countries. In sub-Saharan Africa, for example, it is more than 85%. The vast majority of TFI cases are caused by salpingitis and subsequent pelvic-peritoneal adhesions due to previous or persistent infections. Bacteria climb up the cervix along mucosal surfaces to reach the endometrium and eventually the fallopian tubes. This pathway manifests itself clinically as PID and it has a significant association with subsequent TFI. About 15% of women who have PID end up with TFI, and if the number of PID attacks increases, the likelihood of infertility also increases. However, most women with TFI do not have a clinically diagnosed PID history, but instead have minimally symptomatic or totally asymptomatic salpingitis as a result of an upper genital tract infection. Studying the effects of such infections, especially those without clinical PID, is necessary to explain TFI because of the mostly asymptomatic course of C. trachomatis. To date, evidence has shown that ascending C. trachomatis infection causes irreversible damage to the fallopian tubes, leading to obstructions and thus, infertility. Heat shock protein (HSP60) synthesized by C. trachomatis produces a proinflammatory immune response in the human fallopian tube epithelium, causing scarring and obstruction in the tube [104, 105].

In a series of sero-epidemiological studies examining antibodies to C. trachomatis and chlamydial hsp60 in laparoscopic or HSG-confirmed fallopian tube injury and ectopic pregnancies, previous C. trachomatis infection has been shown to significantly increase tubal infertility in women, regardless of whether it presents clinical symptoms or not. Patients with PID are also more likely to develop infertility later in the presence of a history of C. trachomatis than those without a history of chlamydia [106, 107].

A cohort study, involving 1250 women with demonstrated tubal patency undergoing fertility treatment, examined C. trachomatis seropositivity using IgG1 and IgG3 antibody subtypes [108]. The presence of IgG3 from these two antibody subtypes was shown to be a strong indicator of both failure to conceive and ectopic pregnancy outcomes. This is because IgG3 is related to the inflammatory response in the early phase of the infection and its detection may indicate that either a recent or persistent infection has caused tube damage, although it has not yet caused tubal obstruction [108]. Another study of subfertile women without visible tubal pathology found 33% lower rates of spontaneous pregnancies in the presence of chlamydial antibodies [107, 108]. Coppus et al. suggest that this decline in pregnancy rates may not only be related to the known chronic inflammatory response but also to the fact that persistent CT infection impairs implantation and embryo development due to the autoimmune response to human heat shock proteins [108, 109]. Therefore, chlamydial antibody tests are likely to continue to be an important predictor both in the evaluation of tubal patency and in the evaluation of ectopic pregnancy, intrauterine insemination failure, and embryo and pregnancy loss independent of tubal damage.

In another systematic review investigating the effect of CT infection on female infertility, a positive correlation between infection and infertility was found in 76.47% of the included studies [110]. The study by Menon et al., which included 239 women, showed that up to half of the subfertile women may have CT infection as a concomitant factor [111]. den Heijer et al. also found that CT-positive women were 70% more likely to experience infertility [112]. Davies [113], Ramadhani [114], and Kayiira [115] also showed results that reinforce this relationship in different countries and populations, noting that routine chlamydia screening, along with interventions to prevent initial and recurrent infections, is extremely important to protect women’s long-term reproductive health.

Considering PID, which causes significant adhesions and severe tubal damage, it is easy to conclude that these anatomical causes are detrimental to fertility. However, there are some cases where there is no apparent damage, suggesting that some molecular mechanisms are also involved. Since CT is an intracellular pathogen that disrupts endothelial and tubal muscle structure, it is highly likely to lead to impaired tubal motility and endothelial cilia function. This explains the changes observed in the form of constrictions in the intrauterine and tubal structure after the application of a saline solution to the female reproductive tract during the laparoscopy procedure. Even if CT remains in the female reproductive organs for a short time, it facilitates the settlement of other microorganisms in the area, leads to changes in the structure of the microbiota, and affects gametes and their conjunction by antigenic stimulation [116, 117]. These immunological changes also explain the state of mild endometriosis in women who have had a preliminary CT infection; once an immunological imbalance has occurred, lymphocyte activity becomes insufficient and, which leads to the retention of viable endometrial cells in the pelvic environment [118]. Thus, as well as the endometriosis, CT infection and the mechanical and biochemical damage it creates, induce a hostile environment for gametes in the female reproductive pathway. A significant point here to emphasize is that as age progresses, C. trachomatis is eliminated from the host and can no longer be detected by PCR. Therefore, a more accurate approach would be using long-standing IgG-specific antibodies to detect past infections that may be responsible for infertility [1].