In: Bubaline Theriogenology, Purohit G.N. and Borghese A. (Eds.). International Veterinary Information Service, Ithaca NY (www.ivis.org), Last updated: 26-Jun-2013; A5708.0613 Approaches for Diagnosis of Pregnancy in Female Buffaloes C.H. Pawshe 1 and G. N. Purohit 2 1 Department of Veterinary Obstetrics and Gynecology, Punjabrao Deshmukh Krishi Vishwa Vidhyalaya Akola, Maharashtra, India. 2 Department of Veterinary Gynecology and Obstetrics, College of Veterinary and Animal Science, Rajasthan University of Veterinary and Animal Sciences, Bikaner Rajasthan India. Early identification of non-pregnant dairy buffaloes post-breeding can improve reproductive efficiency by short-cycling the animals and rebreeding during the same breeding season. Thus early and accurate diagnosis of pregnancy seem important to shorten the calving interval in buffaloes [1] that show a profound effect of season on reproduction, coupled with poor estrus expression [2]. The availability of pregnancy diagnosis services appears to be extremely low in some locations [3,4] and at other places the diagnostic approaches seem to be far from perfect. Failure of pregnancy diagnosis leads to slaughter of pregnant buffaloes; with every fourth buffalo slaughtered being pregnant [5] resulting in heavy economic losses. The attributes of an ideal pregnancy diagnostic test include i) high sensitivity (i.e., correctly identify pregnant animals) ii) specificity (i.e., correctly identify non-pregnant animals), iii) inexpensive, iv) simple to conduct under field conditions and v) ability to determine pregnancy status at the time the test is performed. It is desirable to diagnose pregnancy as soon as possible after insemination but without having the diagnosis being confounded by subsequent embryonic mortality. Most direct (clinical) and indirect (laboratory) methods of pregnancy diagnosis include one or more of these attributes, but no test currently available or under development include all the attributes mentioned. Two clinical methods for pregnancy diagnosis in buffaloes have become popular; transrectal palpation is the oldest method and continues to be the most widely used method for pregnancy diagnosis in cattle and buffaloes [6]. However, this approach suffers from the drawback that diagnosis is not accurate before day 45 [7] and early embryonic death may occur possibly due to rough genital manipulation. Although transrectal ultrasonography procedures have gained popularity as a clinical method for pregnancy diagnosis in the buffalo [1,8] providing additional information about the fetus and its surroundings, their specificity and sensitivity is 100% only around Day 30 [1]. A large number of laboratory methods have been developed for pregnancy diagnosis in buffaloes including the detection of circulating hormones, pregnancy-associated molecules or chemical determination of these or other molecules in natural secretions (cervical mucus, urine or milk) from pregnant buffaloes, however, either their detection requires a specialized laboratory setup or their sensitivity and specificity is low limiting their widespread use. In this chapter the authors have classified and discussed methods of pregnancy diagnosis in buffaloes under two subheadings Clinical methods (Non-return to estrus, transrectal palpation, transrectal ultrasonography) and 1. Laboratory methods (Plasma and milk progesterone, plasma estrogen, pregnancy associated proteins and chemical tests). 2. 1. Clinical Methods 1.1. Non-return to Estrus After conception, the embryo inhibits the regression of the corpus luteum (CL) by secreting some molecules that suppress PGF2α secretion thus maintaining high progesterone concentrations that prevent the buffalo from returning to estrus. The non-return to estrus of a bred buffalo 18-24 days after breeding is an assumption by breeders that the animal is pregnant. Alternatively, if the buffalo is not pregnant after breeding, the uterus will secrete PGF2α that will lyse the CL, plasma progesterone concentrations will decrease and the buffalo will return to estrus after 21 to 23 days post-service. The reliability of the method depends on the accuracy of estrus detection in the herd. This approach over-estimates pregnancy by 10-20% [9]. Gestational estrus in a small proportion of pregnant buffaloes also complicates the diagnosis [9- 10]. Poor estrus signs and sub-estrus in some buffaloes [2] lead to their erroneous diagnosis of pregnancy. Moreover, reasons other than pregnancy such as chronic uterine infection or ovarian cysts, may prevent estrus, resulting in buffaloes being erroneously diagnosed as pregnant. Buffaloes bred during the time of increasing day length may fail to return to