Bacterial Isolation and Antibiogram of Uterine Lavage from Repeat Breeder Cows Suffering from Subclinical Endometritis

A total 500 cows were randomly selected from college livestock farm, Kuthuliya and different villages in and around Rewa (M.P.). After recording history all the animals were subjected to gynaeco-clinical examination, Whiteside test and endometrial cytology by cytobrush technique. On the basis of above tests performed all the animals were selected for the study. They were divided into three groups as follows: Normal (n=280), Clinical endometritis (n=80) and Subclinical endometritis (n=140). All the animals were subjected to aseptic collection of uterine fluid by low volume lavage technique. Uterine fluid samples obtained were used for microbial assay and antibiotic sensitivity tests. Among the bacterial isolates Staphylococcus species (36.31%) was highly prevalent. The antibiotic sensitivity of isolates was found to be maximum for ceftriaxone and sulbactum combination 91.67 per cent followed by levofloxacin 89.07, ciprofloxacin 79.69, ceftriaxone 73.43, enrofloxacin 61.45 and gentamicin 56.78 per cent, respectively. It was concluded that Staphylococcus species was highly prevalent bacteria isolated and a combination of ceftriaxone and sulbactum was found to be highly sensitive.

Good fertility in dairy cows is considered as key to economically successful dairy farming. It is widely accepted that uterine disorders in the postpartum period have a negative impact on reproductive performance. Postpartum uterine infection is the major cause of economic loss in dairy cows. A cow that has normal or nearly normal oestrus cycle and bred to 3 or more time with a fertile bull or AI by fertile semen yet fail to conceive is known as repeat breeder. Repeat breeding syndrome results in lowered dairy profit via wastage of semen and increased insemination cost, and reducing fertility. After the postpartum period, repeat breeding is considered one of the most important reproductive disorders in cattle (Yusuf et al., 2010) which affect reproductive efficiency. Generally, non-specific infection of the genitalia is considered to be the main cause of repeated conception failure.
Postpartum uterine infections can delay the endometrial regeneration and disrupt the resumption of ovarian cyclicity which leads to the postponement of first insemination (AI), increase in number of inseminations per conception and thus calving interval is prolonged. Histologically, endometritis is defined as a disruption of the epithelium with presence of inflammatory cells. Subclinical endometritis is defined as an endometrial inflammation occurring 21 days or more after parturition without any clinical signs whereas clinical endometritis is indicated by the presence of purulent/ mucopurulent discharge.
Fertility of cow is affected by many nonspecific and specific pathogens of the genital tract. Uterine bacterial infections or bacterial products also suppress pituitary LH secretion and perturb postpartum ovarian follicle growth and function, which disrupts ovulation in cattle.
A higher percentage of cows (80 to 100%) are found to have bacterial contamination of the uterus in the first 2 weeks postpartum. Many of these bacteria are eliminated during the first 5 weeks after parturition, but the remaining bacteria cause uterine disease in some cows. Common bacteria isolated from cows with uterine infections are Escherichia coli, Arcanobacterium pyogenes (Sheldon et al., 2009).
Major advantages of intrauterine treatment with antibiotics are reduced interval of treatment, rapid recovery rate and improved conception rate but their unselective use has led to development of resistant bacterial strains making further use of such therapy useless. The antibiotics have been use to uterine infections with variable success and the efficacy of antibiotics evaluated from time to time due to continuous emergence of drug resistant bacterial strains (Barman et al., 2013).
Keeping this in view, the present study was planned with an objective to study antibiotic sensitivity of bacterial isolates obtained from normal, clinical and subclinical endometritis of repeat breeder cows.

MATERIALS AND METHODS
A total 500 cows were randomly selected from college livestock farm, Kuthuliya and different villages in and around Rewa (M.P.). After recording history all the animals were subjected to gynaeco-clinical examination, Whiteside test and endometrial cytology by cytobrush technique. On the basis of above tests performed all the animals were selected for the study. They were divided into three groups as follows: Normal (n=280), Clinical endometritis (n=80) and Subclinical endometritis (n=140).
All the animals were subjected to aseptic collection of uterine fluid and endometrial tissue by low volume lavage technique. These samples were collected aseptically in autoclaved Brain Heart infusion (BHI) broth tubes and brought to laboratory in ice boxes. The aseptically collected fluid from all the animals were subjected to culture isolation and identification of bacterial microorganisms (Cruickshank, 1965) as well as for antibiogram as per method described by Barry (1976). Antibiotic sensitivity discs supplied by HI-MEDIA Laboratories Limited, Mumbai (India) were used and tested for their sensitivity with 6 antibiotics using antibiotic sensitivity discs viz., ceftriaxone (30 mcg/disc), levofloxacin (05 mcg/disc), gentamicin (10 mcg/disc), ciprofloxacin (5 mcg/disc), enrofloxacin (10 mcg/disc) and ceftriaxone/ sulbactum (30/15 mcg/disc). The results were interpreted as per the chart furnished by the company.

RESULTS AND DISCUSSION
Different types of bacteria were isolated from uterine fluid samples obtained by low volume uterine lavage technique. All the uterine fluid samples obtained from normal group animals were sterile. All the 80 uterine lavage samples obtained from clinical endometritic cows screened for bacterial isolates were found to be positive. Out of these, 80 samples that were found to be positive for bacterial isolates, 04 (5.00%) samples given mixed bacterial isolates while 76 (95.00%) isolates were of single type. Out of 84 isolates, 35 isolates (41.67%) were of E. coli, 6 (7.14%) of Streptococcus species, 34 (40.48%) of Staphylococcus species and 9 (10.71%) of Bacillus species.
The prevalence of Staphylococcus species isolates as observed in the present study was in accordance with the findings of Behera et al. (2015) and Ahuja et al. (2017). They observed that E. coli and Staphylococcus species was highly prevalent in endometritic buffaloes. Aforesaid findings of present study are in agreement with Samatha et al. (2013) who reported the bacterial isolates in the order of Staphylococcus species (28.12%), E. coli (21.87%) and Pseudomonas (15.62%) in endometritic buffaloes. Above findings are in close accordance with Patel et al. (2009) who reported that Staphylococcus species was most prevalent bacterial isolated in buffaloes.
All the uterine lavage samples found positive (192) for bacterial isolates were subjected to antibiotic sensitivity tests for 6 different antibiotic agents (cefriaxone, levofloxacin, gentamicin, ciprofloxacin, enrofloxacin and ceftriaxone + salbactum). The results of antibiotic sensitivity of the cultures of uterine lavage of cows suffering from clinical and subclinical endometritis.

CONCLUSION
It can be concluded that Staphylococcus species is highly prevalent bacteria causing clinical and sub-clinical endometritis. A combination of ceftriaxone + salbactum was found to be highly sensitive against bacteria causing subclinical and clinical endometritis in postpartum repeat breeder cows.