Infertility

Empirical Treatment of
Low-level Leukocytospermia With
Doxycycline in Male Infertility Patients
Alaa Hamada, Ashok Agarwal, Reecha Sharma, Dan B. French, Ahmed Ragheb, and
Edmund S. Sabanegh, Jr.
OBJECTIVE To design a retrospective study in which infertile men with a seminal leukocyte count of 0.2-1 ⫻ 106
white blood cells [WBC]/mL were given doxycycline to examine the potential benefits of the
treatment on the improvement of semen parameters and natural pregnancy outcomes. It has been
reported that even low-level leukocytospermia (0.2-1.0 ⫻ 106 WBC/mL) could be harmful to
male reproductive function and achievement of pregnancy.
METHODS The records of 223 patients were reviewed and 61 patients were identified with a leukocyte count
of 0.2-1.0 ⫻ 106 WBC/mL. Of the 61 patients, 27 presented before 2006 and were not routinely
treated for this level of leukocytospermia. These patients were our historic control population.
Since 2006, 34 patients met these criteria and received empirical antibiotic therapy (treatment
group).
RESULTS The treatment of low-level leukocytospermia with doxycycline did not show statistically signif-
icant differences in the semen parameters among the treated patients. Although the therapy did
not alter the semen parameters, low-level leukocytospermia resolved in 56% of the treatment
group, well above the spontaneous resolution rate of 25% observed in historic controls. The
natural pregnancy rate among the treatment group (15 of 32 [47%]) was significantly greater than
that among the controls (5 of 25 [20%]). The odds ratio for pregnancy outcome was 3.7 (95%
confidence interval 1.1-11.7; P ⫽ .04).
CONCLUSION Low-level leukocytospermia might have deleterious effects on male fertility, and antibiotic
therapy for such a condition might improve the natural pregnancy rate among infertile
couples. UROLOGY 78: 1320 –1325, 2011. © 2011 Elsevier Inc.

T
he pathologic role and origin of leukocytospermia mal digital rectal examination findings, or there are 2
in male factor infertility and the indications for disordered ejaculate signs, such as leukocytospermia or
treatment remain controversial. Leukocytes are bacteriospermia.2 Infertility due to MAGI is attributed to
present in the semen of both fertile and infertile men, direct microbial damage to the sperm or indirect injury to
even in the absence of florid genital infections. In addi- the sperm through increased activated seminal leukocyte
tion, leukocytospermia might be a presenting feature of release of cytokines and reactive oxygen species (ROS).
prostatitis, seminal vesiculitis, or epididymitis. However, However, the correlation between leukocytospermia
it has been described in 10%-20% of infertile men pre- and bacteriospermia is controversial, particularly in
senting for fertility evaluation without clinical signs of asymptomatic men.3-5 Specific bacterial pathogens and
accessory glands infection, a phenomenon sometimes viral etiology have been identified in only 40%-54.9%
termed “silent accessory gland infection.”1 and 11% of leukocytospermic men, respectively.3,5 To
Male accessory gland infection (MAGI) is defined by complicate the picture, Lackner et al3 identified patho-
the World Health Organization (WHO) when there are genic bacteria in the absence of leukocytospermia in
2 clinical features suggestive of MAGI, such as a history 48.2% of infertile men. Furthermore, the presence of
of urinary tract infection, epididymitis, or sexually trans- bacteriospermia might represent contamination or nor-
mitted disease, and/or abnormal physical signs, such as mal urethral colonization, rather than a real infection,6
thickened or tender epididymis or vas deferens or abnor- and the value of semen culture to identify the pathogenic
bacteria is, therefore, limited. Leukocytospermia in the
From the Center for Reproductive Medicine and Glickman Urological and Kidney absence of infection can be attributed to other causes,
Institute, Cleveland Clinic, Cleveland, Ohio such as smoking, marijuana use, heavy alcohol use, and
Reprint requests: Edmund Sabanegh, Jr., M.D., Department of Urology, Cleveland
Clinic, Q100, 9500 Euclid Avenue, Cleveland, OH 44195. E-mail: sabanee@ccf.org clomiphene therapy. The effect of leukocytospermia on
Submitted: June 1, 2011, accepted (with revisions): August 30, 2011 the semen parameters and pregnancy outcomes is still
1320 © 2011 Elsevier Inc. 0090-4295/11/$36.00
All Rights Reserved doi:10.1016/j.urology.2011.08.062
under debate. Although some studies have demonstrated achievement of pregnancy. We designed a retrospective
a beneficial effect of leukocytospermia on sperm func- study in which infertile men with a leukocyte count of
tion,7 considerable evidence from other studies has 0.2-1 ⫻ 106 WBC/mL were given empirical doxycycline
shown detrimental effects on the semen parameters and therapy to examine the potential benefits of therapy on
sperm function tests, such as the hamster egg fertilization the semen parameters and natural pregnancy outcomes.
test.8-10 Alternatively, no significant relationship be-
tween leukocytospermia and poor semen quality or preg-
MATERIAL AND METHODS
nancy outcomes has been found in other studies.6
The WHO has arbitrarily defined leukocytospermia as Subjects
⬎1 million white blood cells (WBC)/mL of semen,11 and Since September 2006, it has been our practice to treat men
most of the previously mentioned studies have used this presenting for fertility assessment who have a level of seminal
level as the pathologic threshold. This threshold level is leukocytes ⱖ0.2 ⫻ 106 WBC/mL with empirical antibiotic
therapy. The standard therapy is a 3-week course of doxycycline
considered a presumptive indicator of MAGI. However,
100 mg orally taken twice daily. Semen analysis and seminal
seminal leukocyte levels of 0.2-1.0 ⫻ 106 WBC/mL have leukocyte levels are reassessed 3 months after therapy completion.
been associated with significantly greater bacterial counts We conducted a retrospective review of the records of male
than lower leukocyte levels.4 Evidence is growing that patients presenting to our Center for Male Fertility for fertility
low-level leukocytospermia (LLLC) is also detrimental to assessment during the period from September 2006 to February
sperm quality and fertility. Thomas et al9 have demon- 2008 to identify those with seminal leukocyte levels of 0.2-1.0 ⫻
strated that the percentage of normal sperm morphology 106 WBC/mL. The Cleveland Clinic institutional review board
varies inversely with the concentration of seminal leu- approved the present study. A control group was obtained by
kocytes, with a significant difference present using a reviewing the records of consecutive patients with seminal leuko-
cyte levels of 0.2-1.0 ⫻ 106 WBC/mL for a period immediately
threshold value of 0.5 million WBC/mL. Seminal leuko-
preceding the practice of routine treatment of LLLC. The men
cyte levels as low as 0.1 ⫻ 106 WBC/mL have been with the clinical symptoms or signs of MAGI, varicocele, and
associated with elevated ROS levels and increased sperm hormonal disturbances were excluded from the study. Patients
DNA damage.12,13 were also screened with standard urinalysis and excluded if
Because of these findings, recent reports have sug- noted to have pyuria or other evidence of urinary tract infec-
gested that LLLC less than the WHO’s empirical value tion. In each couple, the female partner was fully evaluated, and
might induce sperm damage and support lowering the cutoff female factor infertility was excluded.
value for the diagnosis of leukocytospermia to 0.2 ⫻ 106
leukocyte/mL.3,4,14 This value was considered the cutoff for Semen Analysis
leukocytospermia in our study. Computer-assisted semen analysis was performed in the androl-
The optimal therapy for leukocytospermia remains ogy laboratory using a Hamilton-Thorne-Integrated Visual Op-
controversial. Several investigators have empirically used tical System, version 10, semen analyzer (Hamilton Thorne
antibiotic therapy in infertile men with traditional leu- Biosciences, Beverly, MA). A 5-␮L aliquot was loaded onto a
counting chamber for each semen parameter analyzed and 4-10
kocytospermia with variable success. Some studies found
fields were examined by computed-assisted semen analysis and
no improvement in the semen parameters after antibiotic manually. The semen parameters are reported according to
therapy.15,16 Others have shown a reduction in the sem- 1999 WHO criteria,11 with an additional assessment of mor-
inal leukocyte level and ROS levels after antibiotic ther- phology using the strict criteria of Kruger et al22 and were
apy17 and an increased rate of conception.18 Antibiotic recorded for each patient before and after treatment.
therapy for leukocytospermia has been shown to improve
the pregnancy rates, even among infertile men with other- Seminal Leukocyte Quantification
wise normal semen parameters.19 Furthermore, Berger et To distinguish the WBCs from immature sperm, the seminal
al20 has demonstrated that doxycycline improves the sperm leukocyte levels were assessed with immunoperoxidase staining
fertilizing potential in the hamster egg penetration assay. (Endtz test).23 Peroxidase-positive leukocytes stain brown and
Doxycycline is a commonly prescribed medication for were counted using a Makler’s counting chamber (Sefi Medical,
the treatment of MAGI owing to its excellent penetra- Haifa, Israel) under bright-field microscopy.
tion into prostatic fluid and semen. Although doxycy-
cline inhibits protein synthesis, it appears to be relatively Outcomes Analysis
nontoxic to spermatogenesis. In addition to its wide The pregnancy outcomes were obtained by chart review. If this
antimicrobial spectrum, doxycycline has a direct effect information was not available in the chart, a telephone inter-
on leukocytes. It inhibits leukocyte chemotaxis and view was conducted. Pregnancies resulting from in vitro fertil-
ization and intrauterine insemination were counted as treat-
phagocytosis,21 interferes with the activation of peroxi-
ment failures. Only natural pregnancies occurring within 6
dase system in seminal leukocytes, and hinders the gen- months of treatment were attributed to treatment success. Res-
eration and release of ROS from leukocytes, the most olution of leukocytospermia was defined as a follow-up Endtz
significant source of ROS in semen.8 test measuring ⬍0.2 million WBC/mL.
We have assumed that even LLLC, such as 0.2 ⫻ 106 Statistical analysis was performed using S-plus, version 7.0,
WBC/mL can be detrimental to sperm function and the software (Insightful, Seattle, WA). The paired t test was used to

UROLOGY 78 (6), 2011 1321

Table 1. Mean and standard deviation for control group’s for pregnancy was 3.7 (95% confidence interval 1.1-11.7;
semen parameters before and after antibiotic treatment P ⫽ .04). Pregnancy data were unavailable for 2 patients
Control Group Control Group in each group, and these were not included in the anal-
Before After P ysis. Among the treated patients, the average interval to
Variable Treatment Treatment Value* pregnancy was 2.15 months (range 0-6). Figure 1 dem-
Volume 3.1 ⫾ 1.4 3.0 ⫾ 1.4 .60 onstrates the observed pregnancy rate between the ob-
Sperm 51.0 ⫾ 58.7 48.0 ⫾ 57.2 .77 servation and treatment groups.
concentration Because many patients do not follow-up as planned if
Motility 45.0 ⫾ 24.2 49.0 ⫾ 23.2 .36
Total motile 73.6 ⫾ 105.0 75.6 ⫾ 116.1 .92 conception occurs, only 6 of 15 treated patients who
concentration achieved pregnancy had a follow-up semen analysis. Of
Sperm these 6, 3 (50%) had their leukocytospermia resolved and
morphology 1 each experienced an increase, no change, or decrease
Kruger criteria 3.9 ⫾ 4.5 4.2 ⫾ 4.4 .75 without resolution of the leukocytospermia.
WHO criteria 15.9 ⫾ 9.8 18.1 ⫾ 9.9 .40
Endtz test 0.47 ⫾ 0.16 0.5 ⫾ 0.2 .31 Of all the treated patients, 25 had a follow-up Endtz
test after treatment. Among the controls, 8 of 27 had ⬎1
WHO, World Health Organization.
Data presented as mean ⫾ SD.
Endtz test to allow for the assessment of spontaneous
* P value of mean differences during observation period. change. Resolution of leukocytospermia was noted in
56% (14 of 25) of the treated patients and 25% (2 of 8)
of the controls.
compare the semen parameters between the control and treat-
ment groups and between the pre- and post-treatment param-
eters in the treatment group. Pregnancy outcomes and treat- COMMENT
ment were analyzed using logistic regression analysis.
Associations are expressed as odds ratios and 95% confidence The WHO threshold (1 ⫻ 106 WBC/mL) for leukocy-
intervals. tospermia11 has been challenged by many investigators
owing to the deleterious effects of seminal leukocytes on
the sperm parameters at much lower levels.24,25 Abnor-
RESULTS mal sperm morphology has been observed with leukocy-
Retrospective test reports of 223 infertile men revealed tospermia levels as low as 0.5 ⫻ 106 WBC/mL.9 More-
34 infertile men who had seminal leukocyte levels of over, the presence of any level of leukocytes in the semen
0.2-1.0 ⫻ 106 WBC/mL and received antibiotic therapy. is associated with higher levels of ROS.12 Additionally,
An additional 27 patients were identified who had not ROS levels are significantly higher with seminal leuko-
received antibiotic therapy, representing the historical cyte levels ⬎0.1 ⫻ 106 WBC/mL.13 From the findings in
control group. these reports, we have suggested a level of 0.2 ⫻ 106
The demographic variables were similar between the 2 WBC/mL to diagnose LLLC.
groups. The mean age of the male partners was 35.5 years In the present study, antibiotic therapy (doxycycline)
(range 23-48) among the controls and 34.3 years (range was given to infertile men with a level of 0.2-1 ⫻ 106
23-48) among the treated patients. The female partners WBC/mL. The rationale of prescribing doxycycline was
were similar in age, with a mean age among the partners derived from reports indicating increased bacterial counts
of the controls of 32.2 years (range 24-40) and 32.6 years in men with leukocyte counts ⬎0.2 ⫻ 106 WBC/mL.4
(range 23-42) among the partners of the treated patients. Doxycycline has unique pharmacodynamic actions on
The mean duration of infertility did not differ between seminal leukocytes that interfere with the release of ROS
the controls (24.6 months) and treated patients (26.5 from seminal leukocytes and nonviable sperm.21 To our
months). The semen parameters among the untreated knowledge, this is the first report of prescribing antibiotic
patients and treated patients before any treatment were therapy for LLLC. Lackner et al25 investigated the fertil-
also similar. A remote history of genital tract infection ization and pregnancy rates of assisted reproduction tech-
(minimum 4 years previously) was noted in 4 of 34 nologies using semen samples with leukocytospermia. Al-
patients in the treatment group and 1 of 23 patients in though the overall fertilization rate was similar in both
the control group. patients and control groups, the pregnancy rates were
The semen parameters among the control and treat- almost 50% in the leukocytospermic samples. Therefore,
ment groups during the study period are listed in leukocytospermia does not necessarily have a negative
Tables 1 and 2. In the treatment group, the average effect on outcome when using assisted reproduction tech-
interval between the initiation of antibiotic therapy and nologies.25
follow-up semen analysis was 85.6 days (range 22-188). Previous reports on using antibiotic therapy for leuko-
The semen parameters between the control and treat- cytospermia levels ⬎1 ⫻ 106 WBC/mL in infertile men
ment group before and after therapy are listed in Table 3. have yielded variable results with regard to the improve-
The pregnancy rate among the patients receiving ment of semen parameters and pregnancy rates. Our
doxycycline (15 of 32 [47%]) was significantly greater study failed to identify any improvement in semen pa-
than among the controls (5 of 25 [20%]). The odds ratio rameters (sperm count, motility, morphology). These re-

1322 UROLOGY 78 (6), 2011

Table 2. Treatment group’s semen parameters before and after antibiotic treatment
Treatment Group Treatment Group Treatment
Variable Before Treatment After Treatment Differences or Ratios P Value
Volume 0.45
Mean ⫾ SD 2.9 ⫾ 1.5 2.6 ⫾ 1.2 ⫺0.1 ⫾ 1.1
Median 2.8 2.5 0.2
Range 0.3-6.6 0.5-5.5 ⫺2.5-2.7
Patients (n) 34 24 24
Sperm concentration 0.55
Mean ⫾ SD 45.6 ⫾ 56.8 55.5 ⫾ 68.5 2.2 ⫾ 4.0
Median 26 24.1 1
Range 0.1-260 0-241 0.2-19.2
Patients (n) 34 24 24
Motility 0.83
Mean ⫾ SD 52.2 ⫾ 22.2 51.0 ⫾ 21.0 ⫺2.0 ⫾ 17.8
Median 55.5 48 ⫺3
Range 0-92.0 12.0-88.0 ⫺29-34
Patients (n) 34 23 23
Total motile concentration 0.8
Mean ⫾ SD 77.3 ⫾ 125.7 86.6 ⫾ 148.6 3.3 ⫾ 8.2
Median 29.3 36.1 0.9
Range 0-499 0-597 0.1-37.2
Patients (n) 34 24 24
Sperm morphology
Kruger criteria 0.1
Mean ⫾ SD 4.5 (4.5) 2.4 (2.3) ⫺3.0 (5.5)
Median 4 (0-18) 2 (0-7) ⫺2 (⫺16-2)
Range n ⫽ 19 n ⫽ 17 n⫽9
Patients (n)
WHO criteria 0.15
Mean ⫾ SD 21.8 ⫾ 9.2 15.5 ⫾ 10.5 ⫺13.3 ⫾ 14.0
Median 24 17 ⫺17
Range 6-35 2-33 ⫺25-6
Patients (n) 11 11 4
Endtz test 0.14
Mean ⫾ SD 0.55 ⫾ 0.22 0.39 ⫾ 0.56 ⫺0.20 ⫾ 0.60
Median 0.4 0.09 ⫺0.4
Range 0.2-1.0 0-2 ⫺0.8-1.6
Patients (n) 34 24 24
WHO, World Health Organization.

Table 3. No significant differences found in semen pa-

rameters between control group and treatment group be-
fore and after treatment
P Value (Control vs.
Treatment Group)
Before After
Variable Treatment Treatment
Volume .52 .29
Sperm concentration .72 .66
Motility .23 .75
Total motile concentration .90 .76
Sperm morphology
Kruger criteria .67 .12
WHO criteria .12 .49
Endtz test .10 .34 Figure 1. Improvement in pregnancy rate among treatment
group and controls.

sults are in consistent with other studies that used em- ethoprim/sulfamethoxasole to be effective therapy for
pirical antibiotic therapy in infertile men with traditional traditional leukocytospermia, noting no change in semen
leukocytospermia. Comhaire et al,26 in 1986, treated parameters after treatment. Erel et al15 similarly con-
leukocytospermic patients with doxycycline and observed cluded that antibiotics (doxycycline and/or ceftriaxone)
no differences in the semen parameters. Also, Yanush- did not treat leukocytospermia and noted no change in
polsky et al16 found neither doxycycline nor trim- semen parameters after treatment. Finally, Krisp et al27

UROLOGY 78 (6), 2011 1323

used levofloxacin in their study and reported no improve- rate noted among those whose leukocytospermia had
ment in the semen parameters. However, a meta-analysis resolved. They also reported a 6% pregnancy rate among
by Skau and Folstad28 of 12 studies using different types untreated patients.19 Idriss et al31 also noted a greater
of broad-spectrum antibiotics, such as doxycycline, eryth- pregnancy rate among couples with idiopathic infertility
romycin, and trimethoprim combined with sulfameth- after treatment for 14 days with tetracycline than in
axazole, to treat traditional leukocytospermia, revealed untreated couples. However, 3 other studies failed to
an improvement in semen quality. The improvement show any increase in the pregnancy rate among leukocy-
noted was substantial in sperm morphology and minor in tospermic men receiving antimicrobial therapy.26,32,33
sperm motility and concentration. The main consistent The effects of leukocytospermia on sperm can be at-
criticism of the studies that failed to show any benefit tributed to increased DNA damage and ROS demon-
from antibiotic therapy in regard to the improvement in strated in various studies. The production of ROS causing
semen parameters, including ours, was the brief duration sperm DNA fragmentation12,13 leads to lower pregnancy
of follow-up, which was ⬍3 months. rates after assisted reproduction technology.18 Seminal
Although the mean leukocyte count in our study co- leukocytes are also associated with elevated levels of
hort remained unchanged, we found that leukocytosper- sperm DNA damage,13 particularly in infertile compared
mia resolved (to ⬍0.2 ⫻ 106 WBC/mL) after 3 months of with fertile men.34 Therefore, this noticeable increment
treatment in 56% of the patients. This response is similar in the pregnancy rate among our treatment group with-
to that of Branigan et al,29 who prescribed doxycycline to out any significant improvement in the semen parameters
treat conventional leukocytospermia in men with might have resulted from enhanced sperm function ow-
chronic prostatitis. After 3 months of doxycycline treat- ing to a decrease in the oxidative stress and sperm DNA
ment, 17% of patients showed resolution with therapy damage imposed by the burden of the seminal leukocytes.
alone and 55% of patients with therapy and frequent Vicari17 discovered a significant decrease in the ROS
ejaculations. There were no requirements for ejaculatory level in a group of infertile men with MAGI treated with
frequency among our patients; however, considering they antibiotics, together with a 40% improvement in the
were attempting conception, it is likely that the ejacula- pregnancy rate. Others have demonstrated improved in
tion frequency might have been similar to that of patients vitro sperm penetration of hamster ova in men with
idiopathic male infertility treated with doxycycline for 20
in the study by Branigan et al,29 who were asked to
days.20
ejaculate at least every 3 days.
We acknowledge several limitations in our study. Al-
Despite using different threshold levels for the diagno-
though comparisons with a control group with similar
sis and resolution of leukocytospermia from that in the
characteristics were provided, our study was retrospec-
study by Branigan et al,29 our study found similar resolu-
tive. The impairment in fertility from low levels of sem-
tion rates. This rate was about 56% (14 of 25) among the
inal leukocytes is in part a result of elevated ROS levels
treatment group and favorably exceeded the spontaneous
and increased sperm DNA damage rather than from
resolution rate of 25% (2 of 8) among the controls in our
alterations in traditional semen parameters. Although we
study and 21% among a larger cohort in a separate study
found no difference in the semen parameters, we did not
(our unpublished data). In another study by Branigan et
have data on the ROS levels or DNA damage. Moreover,
al,19 using more traditional threshold values for leukocy- the small sample size might have been a factor in the lack
tospermia, a 68% resolution rate was noted after antibi- of any differences in the semen parameters after treat-
otic therapy. Although a number of studies have demon- ment. Therefore, a larger prospective study incorporating
strated significant resolution of leukocytospermia,17,30 ROS and sperm DNA damage levels before and after
others have failed to show any resolution15,16,27 with treatment is underway. Most studies describing antibiotic
antibiotic therapy. In our study, the percentage of im- therapy for leukocytospermia included treatment of the
provement in the leukocyte count in the treatment group female partner; however, this was not routinely done in
compared with the historic controls can be attributed to our series. No studies have compared treatment of the
doxycycline treatment. A reduction in the bacterial male partner with treatment of both partners to support
counts and products decrease the inciting effects on the this.
seminal leukocytes, resulting in their resolution.
The interesting finding in our study was the improve-
ment in the fertility rate after therapy. In the treatment CONCLUSIONS
group, 47% of patients were able to achieve pregnancy In our study, we examined the effect of treating leuko-
compared with 20% among the untreated patients, with cytospermia at levels from 0.2 ⫻ 106 to 1.0 ⫻ 106
an odds ratio for pregnancy of 3.7 (95% confidence WBC/mL. Although empirical doxycycline therapy does
interval 1.1-11.7, P ⫽ .04). Our results compare favor- not alter the semen parameters, LLLC resolved in more
ably with other reports that examined the natural preg- than one half of our patients, well above the spontaneous
nancy rate in leukocytospermic men treated with antibi- resolution rate. The natural pregnancy rates were signif-
otics. Branigan et al19 found that 37% of treated men icantly improved in the infertile couples in whom the
were able to achieve pregnancy, with a 53% pregnancy men were treated for LLLC. LLLC seemingly has a direct

1324 UROLOGY 78 (6), 2011

deleterious effect on male reproductive potential and 16. Yanushpolsky EH, Politch JA, Hill JA, et al. Antibiotic therapy
perhaps sperm function. Additional studies assessing the and leukocytospermia: a prospective, randomized, controlled study.
Fertil Steril. 1995;63:142-147.
response of sperm function tests, such as ROS levels, 17. Vicari E. Effectiveness and limits of antimicrobial treatment on
DNA damage, and sperm fertilizing potential are needed seminal leukocyte concentration and related reactive oxygen spe-
to better understand the role of LLLC and its treatment cies production in patients with male accessory gland infection.
in male factor infertility. Hum Reprod. 2000;15:2536-2544.
18. Menkveld R. Leukocytospermia. Int Congr Ser. 2004;2004:218-224.
Acknowledgment. To the staff of Cleveland Clinic’s An- 19. Branigan EF, Spadoni LR, Muller CH. Identification and treatment
drology Laboratory for their technical assistance. of leukocytospermia in couples with unexplained infertility. J Re-
prod Med. 1995;40:625-629.
20. Berger RE, Smith WD, Critchlow CW, et al. Improvement in the
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