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2011, Vol. 6 No. 1, Article 79

 

Vaginal Microflora and its Antibiogram in Infertile Bitches

Ambili John, Divya R. Nair, J. R. Sreejith, L. Deepthi, R. Praseeda1 and G. Ajitkumar*

 

Obstetrics and Gynaecology Unit, University Veterinary Hospital,
Faculty of Veterinary and Animal Sciences, Kerala Agricultural University,
Kokkalai, Thrissur, Kerala, INDIA 680 021

1Veterinary Surgeon, Kerala State Animal Husbandry Department

 

*Corresponding Author; e-mail address: ajitvet@yahoo.co.in

 


ABSTRACT

An investigation carried out to ascertain the vaginal microflora in infertile dogs revealed that out of the 17 vaginal discharge samples subjected to culturing, growth could be appreciated in 71 per cent. The organisms identified were E. coli (38.5%), Enterococcus (23.0%), Klebsiella (7.7%), Enterobactor (7.7%), Proteus (7.7%), β-haemolytic Streptococci (7.7%) and Candida (7.7%). Out of the six bacteria tested for antibiotic sensitivity, 83.33% each were found to be sensitive to ofloxacin and gentamicin, 66.67% each to ampicillin/sulbactam, amikacin and cotrimoxazole, 50% each to chloramphenicol and cefotaxime, 33.33% each to netilmicin, cephalexin, cloxacillin and ciprofloxacin and 16.67% each to amoxicillin/clavulanic acid, lincomycin and norfloxacin.

KEY WORDS

Vaginal microflora, infertility, dog.

INTRODUCTION

Allen (1986) reported that most cases of infertility in bitches fall into one of the three main groups such as anoestrus, mistiming of mating and possible interference from non-specific bacteria. Generally, the genital bacterial infections in the infertile bitch are thought to be sub-clinical, but occasionally result in obvious vaginitis, metritis, pyometra or systemic infections. Many aerobic and anaerobic bacteria normally inhabit the vestibule and vagina of the healthy bitch and the bacterial flora is normally mixed. Primary bacterial infection of the canine vagina is less common and occasionally, it can be difficult to distinguish pathogenic bacteria from normal vaginal flora. However, a heavy growth of a uniform bacterium is more suggestive of a pathogen (Freshman, 1991). The two forms of vaginitis recognized in dogs are juvenile or puppy vaginitis and adult-onset vaginitis (Kustritz, 2008). Vaginitis may occur in bitches regardless of age, breed, or reproductive status. Clinically, the most common signs of inflammation of vagina are mucoid to purulent vulval discharge, pollakiuria, scooting and frequent licking of the vulva (Johnson, 1991).
Vaginal cytology is a simple technique that can be used by practitioners to help characterize stages of the reproductive cycle of the bitch or to evaluate certain diseases of the genital tract (Schutte, 1967; Christie et al., 1972; Jones and Joshua, 1988; Arthur et al., 1989; Feldman and Nelson, 1996). Cytologic changes in vaginitis include the presence of non-cornified vaginal epithelial cells and many degenerate or nondegenerate neutrophils. According to Kustritz (2008), the diagnostic tests for vaginitis include a cytologic examination of vaginal epithelial cells and vaginal discharge, vaginal and urine bacterial cultures and antimicrobial sensitivity testing, urinalysis, a digital vaginal examination and vaginoscopy.
Treatment of mild cases of vaginitis in bitches usually involves oral administration of systemic alkalinizers like disodium hydrogen citrate. Johnston et al. (2001) opined that antibiotics should be used if culture of a specimen collected from the anterior vagina reveals heavy growth of a single organism and that systemic therapy is preferred to the topical treatment. Vaginal douches with antibiotic or antiseptic agents are ineffective in flushing out significant amounts of vaginal discharge and may be irritating to the vaginal mucosa, worsening the vaginitis. Knowledge on vaginal microflora and their antibiogram is of paramount importance in the judicious selection of the antibiotic to tackle infertility due to vaginitis. Under these circumstances, an investigation was undertaken to identify the vaginal flora of infertile bitches and the antibiogram.

MATERIALS AND METHODS

Infertile female dogs (n=48) presented for routine exfoliative vaginal cytology as part of the breeding soundness evaluation at University Veterinary Hospital, Kokkalai, Thrissur formed the material for the present investigation. Seventeen bitches of different breeds and age (ranging from 1 to 4 years), diagnosed to be suffering from vaginitis on exfoliative vaginal cytology were selected at random and samples of vaginal discharge collected aseptically from these animals were subjected to routine culture and sensitivity test. Fifteen different antibiotics were employed in the sensitivity test. The details of organisms isolated and the antibiogram data were compiled and analysed.

RESULTS

Out of the 17 vaginal discharge samples collected aseptically and subjected to culturing, growth could be appreciated only in 12 (71%). From among the samples found positive on culture, 13 isolates could be obtained. The organisms identified were E. coli (38.5%), Enterococcus (23%), Klebsiella (7.7%), Enterobactor (7.7%), Proteus (7.7%), β-haemolytic Streptococci (7.7%) and Candida (7.7%) (Fig.1).
The details of the antibiogram pattern obtained for the organisms isolated are summarized in Table.1. Out of the six bacteria tested for antibiotic sensitivity, 83.33% each were found to be sensitive to ofloxacin and gentamicin, 66.67% each to ampicillin/sulbactam, amikacin and cotrimoxazole, 50% each to chloramphenicol and cefotaxime, 33.33% each to netilmicin, cephalexin, cloxacillin and ciprofloxacin and 16.67% each to amoxicillin/clavulanic acid, lincomycin and norfloxacin.

DISCUSSION

An investigation undertaken to identify the vaginal flora of infertile bitches suffering from vaginitis revealed that out of the 17 samples subjected to culturing, 71% were found to be positive and the bacteria identified were E. coli, Enterococcus, Klebsiella, Enterobactor, Proteus and β-haemolytic Streptococci. Candida also could be identified from one of the samples. Of the vaginal culture specimens collected by Bjurstrom (1993) from 78 bitches with vaginitis, 74% were found to be positive for bacterial growth. According to Olson and Mather (1978), the aerobic bacteria isolated from the vaginal tract of normal bitch included E. coli, Staphylococci and Streptococci. The bacteria isolated by Watts et al. (1996) from the vagina of dog during proestrus and oestrus included Alcaligenes faecalis, Corynebacterium spp., Haemophilis spp., E. coli, Streptococcus faecium, Pasteurella spp., Bacteroides spp. and Proteus mirabilis. The present study revealed the presence of both Gram-positive and Gram-negative bacteria in the vagina of bitches suffering from vaginitis.
Perusal of the antibiogram pattern revealed that 83.33% of the bacterial isolates from vaginal discharge of infertile bitches were sensitive to ofloxacin and gentamicin. Ofloxacin, a second generation fluoroquinolone antibiotic that is active against both Gram-positive and Gram-negative bacteria is contra-indicated during pregnancy and lactation. Spontaneous abortions following exposure to fluoroquinolones during pregnancy have been reported in humans (Loebstein et al., 1998). As gentamicin cannot be administered by oral route, on many occasions it may not be the preferred antibiotic for the treatment of vaginitis in infertile bitches. Combination of ampicillin and sulbactam was found to be sensitive to two-third of the bacterial isolates. As animal reproduction studies have failed to demonstrate a risk to the foetus, ampicillin/sulbactam has been assigned to pregnancy category B by the Food and Drug Adminstration (www.fda.gov), USA. Even though amikacin was found to be sensitive to two-third of the isolates, as it has to be administered parentarily, it is not generally selected as the antibiotic of choice in vaginitis. With respect to co-trimoxazole, embryopathic effects such as neural tube defect, congenital heart defect or oral cleft have been reported (Shepard et al., 2002).
In nutshell it may be inferred that when antibiotics are to be used to treat genital tract infections, extreme care should be exercised in the judicious selection of the antibiotic of choice as some of them can cause serious detrimental effects on sperm, fertilized oocytes or early developing embryos. In situations which warrant the use of potent antibiotics, much attention should be paid to ensure proper withdrawal time as there is possibility of persistence of them in the animal’s body at the time of breeding.
Conclusion
An investigation undertaken to identify the vaginal flora of infertile bitches and their antibiogram revealed E. coli, Enterococcus, Klebsiella, Enterobactor, Proteus and β-haemolytic Streptococci as the predominant organisms and that 83.33% of the isolates were found to be sensitive to ofloxacin and gentamicin.

ACKNOWLEDGEMENTS

Thanks are due to Dr. P. C. Alex, Professor and Head, University Veterinary Hospital, Kokkalai, Thrissur for the facilities provided and Dr. E. Nanu, Dean, Faculty of Veterinary and Animal Sciences, Kerala Agricultural University, Mannuthy for granting permission to publish this paper.

REFERENCES

  1. Allen, W.E. (1986). Infertility in the bitch. In Pract. 1: 22-26

  2. Arthur, G.H., Noakes, D.E. and Pearson, H. (1989). Veterinary Reproduction and Obstetrics. Sixth edition. ELBS, Bailliere Tindall, London, pp. 26-30, 487-505

  3. Bjurstrom, L. (1993). Aerobic bacteria occurring in the vagina of bitches with reproductive disorders. Acta Vet. Scand. 34: 29-34

  4. Christie, D.W. Bailey, J.B. and Bell, E.T. (1972). Classification of cell types in vaginal smears during the canine oestrous cycle. Br. Vet. J. 128: 301-309

  5. Feldman, E.C. and Nelson, R.W. (1996). Ovarian cycle and vaginal cytology. In: Canine and Feline Endocrinology and Reproduction. Second edition. W. B. Saunders Co., Philadelphia, pp. 526-545, 633-638

  6. Freshman, J.L. (1991). Clinical approach to infertility in the cycling bitch. Vet Clin N Am: Small Anim Pract 21:427-435

  7. Johnson, C.A. (1991). Diagnosis and treatment of chronic vaginitis in the bitch. Vet Clin N Am: Small Anim Pract 21:522-531

  8. Johnston, S.D., Kustritz, M.V.R. and Olson, P.N.S. (2001). Canine and Feline Theriogenology. Saunders, Philadelphia. 592p

  9. Jones, D.E. and Joshua, J.O. (1988). Reproductive Clinical Problems in the Dog. Second edition. Wright Publishers, London, pp. 33-38

  10. Kustritz, M. V. R. (2008). Vaginitis in dogs: A simple approach to a complex condition. Vet. Med. 103:562-567

  11. Loebstein, R., Addis, A. and Ho, E. (1998). Pregnancy outcome following gestational exposure to fluoroquinolones: a multicenter prospective controlled study. Antimicrob. Agents Chemother. 42: 1336–1339

  12. Olson, P.N.S. and Mather, E.C. (1978). Canine vaginal and uterine bacterial flora. J. Am. Vet. Med. Assoc. 172: 708

  13. Schutte, A.P. (1967). Canine vaginal cytology-I. Technique and cytological morphology. J. Small Anim. Pract. 8: 301-306

  14. Shepard, T.H., Brent, R.L., Friedman J.M., Jones K.L., Miller R.K., Moore, C.A. and Polifka, J.E. (2002). Update on New Developments in the Study of Human Teratogens. Teratology 65:153–161

  15. Watts, J. R., Wright, P. J. and Whithear, K. C. (1996). Uterine, cervical and vaginal microflora of the normal bitch throughout the reproductive cycle. J. Small Anim. Pract. 37:54-60


TABLES

Table 1: Details of organisms isolated from vaginal discharge of infertile bitches and their antibiogram pattern

Serial No

Organism isolated

Antibiotic to which sensitive

Zone of inhibition obtained (mm)*

1

Escherichia coli (n=5)

Ampicillin/Sulbactam

Ofloxacin

Chloramphenicol

Cefotaxime

Piperacillin

Gentamicin

Amikacin

Clotrimoxazole

Netimicin

24,22

24,19

24,24,20

24

22

22,20,20

22,20

22,21

20

2

Enterococcus spp. (n=3)

Cefotaxime

Lincomycin

Ciprofloxacin

Ofloxacin

Amoxycillin/Clavulanic acid

Cephalexin

Ampicillin/Sulbactam

Gentamicin

Amikacin

Cloxacillin

Netilmicin

28

24

24

22

22

22,22

20,20

20

20,20

20

20

3

Klebsiella spp. (n=1)

Clotrimoxazole

Chloramphenicol

Ofloxacin

Norfloxacin

Gentamicin

24

24

24

22

22

4

Enterobacter spp. (n=1)

Ampicillin/Sulbactam

Cotrimoxazole

Gentamicin

Ofloxacin

Amikacin

24

24

22

22

22

5

Proteus spp. (n=1)

Clotrimoxazole

Ofloxacin

Ampicillin/Sulbactam

Chloramphenicol

Gentamicin

22

20

18

18

16

6

β-haemolytic Streptococci (n=1)

Cefotaxime

Cloxacillin

Ciprofloxacin

Cephalexin

Amikacin

24

22

22

20

20

Zone of inhibition (mm) for the antibiotic to become sensitive: Amikacin-17, Amoxicillin/Clavulanic acid-18, Ampicillin/Sulbactam-15, Cefotaxime-23, Cephalexin-18, Chloramphenicol-18, Ciprofloxacin-21, Clotrimoxazole-16, Gentamicin-15, Linkomycin-18, Netilmicin-15, Norfloxacin-17, Ofloxacin-16.

 

FIGURES

Figure 1. Percentage of various isolates obtained on culturing vaginal discharge from infertile bitches

 

 


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