CONSTRAINTS LIMITING THE EFFICIENCY OF ARTIFICIAL INSEMINATION OF CATTLE IN BANGLADESH

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1 XA00320 CONSTRAINTS LIMITING THE EFFICIENCY OF ARTIFICIAL INSEMINATION OF CATTLE IN BANGLADESH M. SHAMSUDDIN, M.M.U. BHUIYAN, T.K. SIKDER, A.H. SUGULLE, P.K. CHANDA, M.G.S. ALAM Department of Surgery and Obstetrcs, Faculty of Veternary Scence, Bangladesh Agrcultural Unversty, Mymensngh, Bangladesh D. GALLOWAY Department of Veternary Scence, Unversty of Melbourne, Werrbee, Australa Abstract CONSTRAINTS LIMITING THE EFFICIENCY OF ARTIFICIAL INSEMINATION OF CATTLE IN BANGLADESH. The am of the present study was to dentfy the factors that nfluence postpartum ntervals to frst detected luteal actvty, frst servce and to concepton, and the concepton rates of cows n the artfcal nsemnaton (AI) programme n Bangladesh. A baselne survey (nvestgatons, 2 and 3) was made on 444 mlkng cows of varous breeds presented for the frst postpartum nsemnaton by 43 farmers lvng at 82 vllages/regons n Mymensngh Dstrct to 6 AI centres and sub-centres. Each cow was then examned three tmes after each AI untl she stopped returnng to oestrus. Sxty to 20 days after the last AI, the cows were examned per rectum to confrm the pregnancy. Mlk progesterone data on Day 0 and Day 2-24 contrbuted to a clear dagnoss wth respect to pregnancy n 82.5% cows ndcatng a possble use of ths progesterone assay schedule for pregnancy dagnoss n AI programmes. The ntervals to frst servce and to concepton vared from 3 to 427 days (medan = 84; n = 444) and 40 to 426 days (medan = 84; n = 232) respectvely, and concepton rate from 32-58% (average 46.2%; n = 444). Prolongaton of weanng age of calves resulted n long ntervals to frst servce and to concepton (P <0.00); weanng age vared from 6 to 9 months (medan = 0). Cows wth body condton score (BCS; -5 scale) of 3 or more and cows calved durng uly to September had shorter ntervals to frst servce and concepton than those wth BCS less than 3 and those calved durng March. The concepton rate was nfluenced by cattle rearng systems (ntensve vs. extensve), purpose of rearng cows (dary vs. dary + draught), BCS and mlk producton (P <0.05). The degrees of vulvar swellng, nature of gental dscharge, toncty of uterus, and nterval between oestrus and AI had sgnfcant effects on the concepton rate. Bulls classfed as good and poor on the bass of semen evaluaton data dffered wth respect to the concepton rate n AI (P O.00); ths ndcates a way of dscrmnatng to some extend between bulls lkely to have hgher or lower fertlty. In Investgaton 4, mlk progesterone was montored two tmes n a month wth a 0-day nterval n 88 cows. The samples were taken between 0 days after calvng and the frst detected oestrus followed by two more samples 0 days apart. The proporton of cows accurately detected n oestrus was 30%. Another 30% were stated to be n oestrus when they were not (false postve) and 40% were not detected when they were n oestrus (false negatve). The ntervals between calvng and oestrus, and luteal actvty were (medan = 20, n = 82) and (medan =, n = 64) days, respectvely. The BCS at calvng and at the ntaton of luteal actvty nfluenced the nterval between calvng and luteal actvty (P <0.05). Cows suckled twce daly ntated luteal actvty earler than ther counterparts suckled several tmes daly (P <0.05). Investgaton 5 demonstrated a reducton n sperm motlty (P <0.05) due to dluton of fresh semen, and chllng, freezng, storng and transportaton of frozen semen. The prolonged postpartum nterval between calvng and concepton and low concepton rate are the maor constrants lmtng the success of AI for cattle development n Bangladesh. The nutrtonal condton of the cow at calvng and thereafter, weanng age of calves, frequency of sucklng, cattle rearng system, accuracy of heat detecton, nterval between oestrus and AI, the oestrus sgns and semen qualty are the mportant determnants of the nterval to concepton and concepton rate.. INTRODUCTION Many farms n Bangladesh are so small that only one cow can be kept. Cows are tethered n a stable or on avalable grazng land. They are used for draught work as well as mlk producton and weanng s not controlled. These management practces promote the occurrence of postpartum

2 anoestrus and lmt behavoural manfestatons of oestrus [, 2, 3]. Most cows are Bos ndcus whch show weak oestrus sgns for a shorter duraton than Bos taunts cows [4, 5, 6]. Detecton of oestrus and of the return to oestrus after unsuccessful artfcal nsemnaton (AI) s clearly dffcult under those condtons and neffcences have been documented [7]. Tradtonally pregnancy dagnoss s not carred out as part of the AI programmes. Veternary servces are not always avalable. There s therefore a need to ntroduce other methods to establsh the status of cows wth respect to cyclcty and pregnancy n assocaton wth AI programmes. The results of AI n Bangladesh are poor wth a concepton rate of 37% after nsemnatng wth chlled semen compared wth 6% after natural servces [8]. Yet AI has the potental to mprove productvty of cows on small farms [9] f hgh reproductve effcency can be acheved. The ams of the present work were (a) to use the radommunoassay of progesterone n mlk as a dagnostc tool to survey some aspects of reproducton n Bangladesh cows and (b) to defne the maor factors lmtng reproductve effcency n smallholder farms usng AI n Bangladesh. 2. MATERIALS AND METHODS 2.. Baselne survey 2... Descrpton of proect area, AI actvtes, farms and cows The nvestgaton was carred out on 444 mlkng cows of varous breeds presented for frst postpartum AI. Most were Bos ndcus and others were crosses of Bos ndcus wth Holsten-Fresan and a mxture of natve breeds. The cows belonged to 43 farmers lvng at 82 vllages/regons n Mymensngh Dstrct - one of the 64 admnstratve unts n Bangladesh. The feld work took place between August 995 and une 997. The farmers' land totalled between 0 and 40 (medan = 0.6) hectares wth -70 (medan = 2.0) breedable cows. The farmers presented ther cow after 5 to 32 (medan = 7) h from oestrus detecton, walkng 0 to 7 (medan = 2.0) km, for nsemnaton by any of the 7 nsemnators; the 0 km ndcates the stuaton where nsemnatons were done n the farms. The cows were beng mlked to 2 (medan =.0) tmes per day wth ther calves at foot. The cows were 4 to 8 (medan = 7.4) years old, and ther party ranged from to 2 (medan = 2), body weght measured from 03 to 480 (medan = 96) kg, body condton scored from to 5 (medan = 3) and mlk producton vared from 0.3 to 6.0 (medan = 2) L per day. Controlled weanng was not practsed, therefore, the tme of spontaneous weanng was recorded. The AI techncans receved 2.5 to 2 (medan = 2) months formal tranng and had 4 to 36 (medan = 6.9) years of practcal experence wth AI n cows. They do between 50 to 250 (medan = 75) nsemnatons per month. Semen used belonged to 46 batches wth sperm motlty 65 to 75% and 29 to 70% before and after processng, respectvely, from 26 bulls representng 7 breeds. Frozen semen n 0.25 ml French straws was used for 254 nsemnatons and 248 nsemnatons were made wth chlled semen. In the cases of chlled semen, ndvdual cow doses were ether ml (n = 95) or 2 ml (n = 53) Investgaton. Use of radommunoassay of progesterone n mlk to survey some reproductve characterstcs of cows n an AI programme To analyse progesterone concentraton, mlk samples were collected n vals contanng sodum azde tablets (8 mg; Merk, Darmstadt, Germany) for preservaton. Mlkng cows presented for frst postpartum nsemnaton were ncluded n ths study. The Day 0 (day of nsemnaton) mlk samples were collected by the AI techncan mmedately after AI. The research personnel pcked-up the samples wthn two days after collecton. The Day 9 3 and Day 2-24 mlk samples were collected by research personnel drectly from the cow at a farm vst. The mlk samples were centrfuged, skmmed mlk was separated and stored at -20 C untl analysed. Progesterone concentraton n mlk was determned by usng sold phase radommunoassay (RIA) kts suppled by the FAO/IAEA Venna. The ntra-assay co-effcent of varance (CV) wth nternal qualty control (IQC) samples vared from 8.0 to 5.6% (0 assays, each wth 0 replcates) and the nter-assays CVs were 6.6% and 8.4% for begnnng and end IQC samples, respectvely (number of assays = 0). 0

3 The data on mlk progesterone concentratons at Day 0, Day 9-3 and Day 2-24 were compared wth the results of per rectum pregnancy dagnoss. Progesterone data based on two samples (Day 0 and Day 9-3) were used to examne the effcency of oestrus detecton. Mlk progesterone concentraton on the day of AI was used to determne the proportons of AI done n the luteal phase of the cow Investgaton 2. Reproductve effcency of cows n an AI programme The nterval from calvng to frst servce, calvng to concepton and the frst servce concepton rate were used as ndces of reproductve effcency. Four hundred and forty four mlkng cows presented for frst postpartum AI were ncluded n ths study. After each AI, the techncan flled n the prescrbed form to record nformaton on nterval from oestrus to AI, oestrous behavour reported by the farmers, degrees of vulvar swellng and uterne tone, and the characterstcs of gental dscharge. The nformaton about farms and cows was recorded between Day 9 and Day 3 at a farm vst. The age of the cow was determned by dental examnaton and the partes were confrmed by questonng the farmers. The body weght of the cow was estmated by usng a standardsed tape (Swedsh Assocaton for Lvestock Breedng and Producton, Esklstuna, Sweden). The nutrtonal condton of the cow was scored (-5) followng modfcaton of the methods descrbed by Ncholson and Butterworth [0]. Between Day 60 and Day 20 after the last recorded AI, all cows were examned per rectum for confrmaton of pregnancy; a mlk sample was always collected mmedately before rectal palpaton f the cow was not dry at that tme Investgaton 3. Evaluaton of semen and classfcaton of bulls Three nsemnaton doses from dfferent batches of ndvdual bulls were examned for sperm motlty, concentraton and proporton of normal spermatozoa wth respect to the acrosome, mdpece and tal [, 2]. To grade the bulls as good or poor, crtera were set based on semen evaluaton data. For chlled semen, f the nsemnaton dose of a bull contaned >50% sperm motlty, >7.5 x 0 6 total motle spermatozoa, >70% normal spermatozoa wth regard to acrosome, mdpece and tal, and >80% spermatozoa wth normal head morphology then the bull was regarded as good. For frozen semen, the correspondng crtera were >30% motlty, >7.5 x 0 6 total motle spermatozoa per nsemnaton dose, >65% normal spermatozoa wth regard to acrosome, mdpece and tal, and >80% spermatozoa wth normal head morphology. If semen characterstcs from a bull dd not meet one or more of these crtera he was classfed as poor. The values set n the crtera were the mean of data on 3 semen samples of ndvdual bulls mnus standard devaton. The classfcaton of bulls was completed before analysng fertlty data Investgaton 4: Accuracy of oestrus detecton studed by usng radommunoassay of progesterone n mlk In the area and AI programmes descrbed n the secton 2..., the feld work for ths nvestgaton was carred out durng uly 997 and December 998. Eghty-eght cows n 58 farms were regstered wthn week after calvng and relevant nformaton wth regard to the farm and cattle was recorded. For ndvdual cows, nformaton was collected on age, breed, party, last calvng date, body weght and body condton score at calvng, occurrence of oestrus, and the occurrence of any post parturent dsorders. Cows that requred maor assstance durng parturton and/or that were dagnosed wth perparturent dsorders lke retaned placenta, puerperal metrts, postpartum haemorrage, prolapse of the gental tract and mlk fever were not ncluded n the nvestgaton. We requested the farmers to report the date and sgns of oestrus but there was no dscusson between the proect personnel and farmers on the sgns of oestrus and ther relevance to concepton. The research personnel collected mlk samples from ndvdual cows twce a month at a 0-day nterval between 0 days postpartum and the frst report of oestrus by the farmer. Two more samples were collected at 0 days nterval, after the occurrence of oestrus. Durng mlk samplng, the research personnel scored the cow for body condton, measured her for body weght, asked the farmer about the occurrence of oestrus and examned the cow for the presence of any dry or fresh dscharge adherng to the perneum. The mlk samples were processed as n the case of Investgaton.

4 2.3. Investgaton 5. Evaluaton of frozen semen from producton to nsemnaton Fve crossbred bulls were examned for breedng soundness accordng to the methods descrbed elsewhere and they were regarded as clncally normal [3]. Semen from the bulls was sampled mmedately after collecton, ntal dluton, coolng down to +4 C, and after storage at -96 C for day, 7 days and 3 months n the Central Cattle Breedng Staton (CCBS), Savar, Dhaka. The same batches of semen were transported to the Dstrct AI Centre, Mymensngh, and then samples were collected after 7 days, 3 months and mmedately before nsemnaton. From the Dstrct AI Centre semen was transported to the Sub Centre, Fulbara and sampled there wthn 7 days after transportaton and mmedately before nsemnaton. The experment was repeated 3 tmes. The semen samples were examned for sperm motlty and proportons of normal spermatozoa wth respect to acrosome, mdpece and tal Analyss of data The data for base lne survey (Investgatons, 2 and 3) were entered n a database applcaton (artfcal nsemnaton data applcaton; AIDA), processed n Excel worksheets and analyses were made usng lnear regresson or a general lnear model [4]. To determne the factors affectng the ntervals from calvng to frst servce and to concepton, the followng regresson model was used: INT = a + blxl + b2x2 + b3x3 + b4x4 + b5x5 + b6x6 + b7x7 + b8x8 + b9x9 + bloxlo + bllxll + e where: INT = Logs of ntervals from calvng to frst servce and to concepton (days) a = constant xl = total land (hectare) held by the farmers x2 = number of breedable cows per farm (counts) x3 = dstance between the AI centres and the farms (km) x4 = age of calves at weanng (months) x5 = age of the cow (years) x6 = party of the cows (counts) x7 = month of calvng (anuary = ; December =2) x8 = month of dong AI (anuary = ; December =2) x9 = body weght (BW) of the cow at AI (kg) xlo = body condton scores (BCS) of the cow at AI (-5 scale) x = mlk produced by the cow at AI (kg) e = error term To determne the effects of grouped and categorcal varables on the ntervals from calvng to frst servce or to concepton a general lnear model was used: INT = n + blxl + b2x2 + b3x3 + b4x4 + b5x5 + b6x6 + b7x7 + b8x8 + b9x9 + bloxlo + bllxll+e where: INT= Logs of ntervals from calvng to frst servce and to concepton (days) u = general mean xl = cattle rearng systems (extensve vs. ntensve) x2 = purpose of rearng cattle (dary vs. dary + draught) x3 = frequency of sucklng (once or twce daly vs. several tmes a day) x4 = feedng system (concentrate added vs. no concentrate added) x5 = breed of cows (crossbred Fresan, crossbred Sahwal, local) x6 = month of calvng (anuary = ; December =2) x7 = month of dong AI (anuary = ; December = 2) x8 = BCS of the cow at AI (Groups = BCS.0-2.0, 2 = BCS , 3 = BCS ) e = error term 2

5 Ch-square test was used to determne the effect on the concepton rate of the followng factors: () Purpose of rearng cattle: Dary vs. dary + draught (2) Frequency of sucklng: Once or twce daly vs. several tmes a day (3) Breed of cows: Crossbred Fresan, crossbred Sahwal, local (4) BCS: Groups = BCS.0-2.0, 2 = BCS , 3 = BCS (5) Daly mlk producton (kg): Groups = < kg, 2 = >l-2 kg, 3 = >2-4 kg, 4 = >4-6 kg (6) Cattle rearng system: ntensve vs. extensve (7) Oestrus-to-AI nterval: -8 h vs h (8) Sgns of oestrus: Standng to be mounted or mountng others, bellowng, gental dscharge or restlessness (9) Degrees of vulvar swellng: Marked vs. slght or mperceptble (0) Nature of gental dscharge: Clear mucus, turbd, none or purulent () Toncty of the uterus at AI: Marked vs. slght or mperceptble (2) Breed of bulls: Fresan, Fresan x local, Sahwal x others (3) Grades of bulls: Good vs. poor (4) Sources of semen: Imported vs. locally produced (5) Types of semen: Frozen vs. chlled (6) Total spermatozoa per cow dose: <5 mllon, 6-20 mllon, 2-25 mllon, mllon (7) AI techncans: n = 7 In Investgaton 4, the followng model was used to analyse the data: INT= n + blxl + b2x2 + b3x3 + b4x4 + b5x5 + b6x6 + b7x7 + b8x8 + b9x9 + e where: INT= log of nterval between calvng and the frst detected luteal actvty (days) LX = general mean xl = cattle rearng system (extensve vs. ntensve) x2 = purpose of rearng cows (dary vs. dary + draught) x3 = feedng system (concentrate fed vs. no concentrate fed) x4 = breed of cows (crossbred Fresan, crossbred Sahwal, local) x5 = BW at calvng (Groups = <200 kg, 2 = kg, 3 = kg, 4 = >30 kg) x6 = BCS at calvng (Groups = BCS.0-2.0, 2 = BCS 2.5, 3 = BCS 3.0, 4 = BCS ) x7 = BW at the frst detected luteal actvty (Groups = <200 kg, 2 = kg, 3 = kg, 4 = >30 kg) x8 = BCS at the frst detected luteal actvty (Groups = BCS.0-2.0, 2 = BCS 2.5, 3 = BCS 3.0,4 = BCS ) x9 = frequency of sucklng (once or twce daly vs. several tmes a day) e = error term The relatonshp between the postpartum perod to ntal luteal actvty and the estmated frst rse n concentraton of mlk progesterone was tested by usng Pearson correlaton. The estmated rse concentraton refers to the mean of the frst detected progesterone rse (> nmol/l) and the progesterone concentratons n samples mmedately before and after the frst rse. ANOVA was used to test the effect of the accuracy of oestrus detecton on the postpartum nterval to oestrus (false postve cases were deleted) and on the estmated progesterone concentraton at the frst peak. Unless otherwse ndcated, the data are presented as medan and range owng to hgh ndvdual varatons. Only the factors that tended (P = 0.0) to or sgnfcantly nfluenced the dependent varables are presented n the Fgures and Tables. In Investgaton 5, repeated measures analyss of varance wth dfferent contrast was used to analyse data on sperm motlty and proporton of normal spermatozoa (wth regard to the arosome, mdpece and tal) obtaned from fresh eaculates, and from semen after dluton, chllng and 3

6 equlbraton, freezng, storage at CCBS for 7 days and 3 months, transportaton to the Dstrct AI Centre and storng there for 3 months, thawng and preparaton for nsemnaton at the Dstrct AI centre, transportaton to an AI Sub Centre, and after thawng and preparaton for nsemnaton at the Sub Centre [4]. In general, the data were log-transformed to near normalty except the proporton values whch were transformed by arcsne transformaton. 3. RESULTS 3.. Investgaton. Use of radommunoassay (RIA) of progesterone n mlk to survey some reproductve characterstcs of cows n an AI programme The nterpretaton of progesterone data based on three samples (Day 0, Day 9-3 and Day 2-24) wth respect to pregnancy results s shown n the Table I. Mlk progesterone data gave a clear nterpretaton on 82.5% cows (n = 360) about ther pregnancy status when comparson was made wth the data of per rectum pregnancy dagnoss at Day None of the 75 cows wth a progesterone profle of low (<.0 nmol/l), hgh (>3 nmol/l) and low on Day 0, Day 9-3 and Day 2-24, respectvely, were found pregnant at rectal palpaton. Twenty-seven cows (7.5%) had a progesterone profle of low hgh and hgh on Day 0, Day 9-3 and Day 2-24, respectvely, but were not pregnant at rectal examnaton. Two nsemnatons were made n pregnant cows. All cows confrmed pregnant at rectal palpaton had a hgh level of progesterone n mlk at the day of pregnancy dagnoss. Eghty one percent of cows (n = 478) had a progesterone profle of low and hgh on Day 0 and Day 9-3, respectvely, ndcatng AI done not at luteal phase or durng ovaran acyclcty (Table II). Ffty-nne cows (2.3%) receved AI at an ncorrect tme as evdent from a devant progesterone profle. Twenty- seven of 506 (5.3%) cows were nsemnated when (Day 0) they had hgh to ntermedate level of progesterone n mlk. TABLE I. MILK PROGESTERONE DATA FROM THREE SAMPLES AND INTERPRETATION IN RELATION TO MANUAL PREGNANCY DIAGNOSIS DayO (day of AI) ' Day 9-3 Day 2-24 Number of Rectal palpaton results; nterpretaton 2 Hgh Hgh cases 202(56.) Pregnant 'ntermedate Hgh 7 (.9) Pregnant; RIA problem, bologcal varatons Hgh 75 (20.8) Non-pregnant; fertlsaton falure, early embryonc death, post AI anoestrus Intermedate 7 (.9) Non-pregnant; fertlsaton falure, short luteal phase, RIA problem, bologcal varaton 6(.7) Non-pregnant; AI at ncorrect tme, post AI anoestrus Intermedate /nter- /hgh medate/hgh Clear nterpretaton 297 (82.5) Hgh Hgh 27 (7.5) Hgh Hgh Hgh Intermedate Hgh 2(0.6) 4(.) Hgh Intermedate 20 (5.6) Intermedate 2 (0.6) Non-pregnant; late embryonc death (>6 days) luteal cyst, persstent corpus luteum (CL) Pregnant; AI on pregnant anmal Non-pregnant; RIA problem, bologcal varaton, late embryonc death, persstent CL Non-pregnant; fertlsaton falure, late embryonc death RIA problem, bologcal varaton Non-pregnant; AI n anoestrous cow, RIA problem Non-pregnant; AI at ncorrect tme, luteal cyst, persstent CL Intermedate/ /nter- Intermedate 8 (2.2) Hgh medate/hgh /hgh Total number of observatons 360 = <.0 nmol/l, 2 Hgh = >3.0 nmol/l, ntermedate = between.0 and 3.0 nmol/l 4

7 TABLE II. MILK PROGESTERONE DATA ON THE DAY OF SERVICE AND ON DAY 9-3 WITH RESPECT TO THE ACCURACY OF OESTRUS DETECTION Day 0 (days of AI) Day 9-3 Number of cases f0/ - "Hgh 387(8.0) Hgh Hgh Hgh 47 (9.8) 6(.3) 6(.3) Interpretatons Progesterone concentraton wthn negatve range on Day 0 and wthn postve range on Day 9-3 ndcates an ovulatory cycle-accurate oestrus detecton. Progesterone concentraton wthn negatve range on both days ndcates anoestrus, anovulaton, or short luteal phase-naccurate oestrus detecton. Progesterone concentraton wthn postve range on both days ndcates AI n pregnant anmals or n anmals wth luteal cystnaccurate oestrus detecton. Progesterone concentraton wthn postve range on Day 0 and wthn negatve range on Day 9-3 ndcates that AI was performed durng luteal phase-naccurate oestrus detecton. Total occurrence 478 Total naccurate oestrus detecton 59(2.3) ] = <.0 nmol/l, 2 Hgh = >3.0 nmol/l, 3 Intermedate = between.0 and 3.0 nmol/l. Thrty-two (6.7%) servces were made n cows wth an ntermedate level of mlk progesterone on Day 0, Day 9-3 or on both occasons Investgaton 2. Reproductve effcency of cows n an AI programme The ntervals from calvng to frst servce and to concepton, and the mean frst servce concepton rate were (medan = 84; n = 444) and (medan = 84; n = 232) days and 46.2% (n = 444), respectvely. The results for the ntervals between calvng and frst servce, and concepton rate are shown n Fgures to 4. The effect of age of cows on the ntervals from calvng to frst servce and to concepton was not sgnfcant (P >0.0). The effect of body condton at AI was sgnfcant on the postpartum ntervals to the frst servce (P <0.00) and to concepton. (P <0.05), and on the concepton rate (P <0.00). Cows wth BCS 3.5 or more took shorter ntervals to receve frst postpartum servce and to conceve than those wth BCS 3 or less (Fgure A and IB; P <0.0). The concepton rates of cows wth BCS.0-2.0, and were 36%, 46% and 64%, respectvely. The crossbred Sahwal cows had a shorter calvng to frst AI nterval than the crossbred Fresan and the pure local cows (Fgure C; P <0.05). The crossbred Sahwal cows had a shorter postpartum nterval to concepton than the crossbred Fresan and local cows (Fgure ID); n both cases, the dfferences between crossbred Sahwal and crossbred Fresan were sgnfcant (P <0.05). The dfferences between breeds n concepton rate was not sgnfcant (P = 7; 54% n crossbred Fresan, 48% n crossbred Sahwal and 43% n local cows). The postpartum ntervals to frst servce and to concepton ncreased wth ncreasng age of the calves at weanng (Fgure 2A and 2B; P <0.00). The relatonshp between daly mlk yeld at frst AI and the ntervals to the frst postpartum AI was not sgnfcant (Fgure 2C and 2D, P >0.0). The effect of mlk producton on the concepton rate was sgnfcant (P <0.05). Cows producng <.0, >- 2, >2-4 and >4 6 kg mlk daly had concepton rates of 36%, 45%, 50% and 58%, respectvely. The dary cows took shorter tmes after calvng to receve frst servce (medan = 74, days; n = 35) and to acheve concepton (medan =8, 40^05 days; n = 75) than the dary +draught cows; the medan nterval from calvng to frst AI was 24 (53-426; n = 29) days, and to concepton 5

8 " " n=6 n=255 n=73 BCS of the cow at AI (-5 scale) " 300 " " - ' ',,,,, n=49 n=34 n=48 BCS of the cow at AI (-5 scale) " " Breed of the cow l=crossbred Fresan, 2=Crossbred Sahwal, 3=Natve Zebu Breed of the cow l=crossbred Fresan, 2=Crossbred Sahwal, 3=Natve Zebu FIG. (A-D). Effect of body condton score and breed of cows on the ntervals from calvng to frst AI and to concepton was 233 (53^26; n = 57) days (P <0.05). Smlarly, the concepton rate was hgher (P <0.05) n dary cows (50%) than that n dary + draught cows (38%). Cows managed ntensvely tended to conceve at a hgher (P = 0.05) rate (53%; n = 56) than those reared extensvely (43%; n = 288). However, the effects of rearng system on the ntervals between calvng and frst servce (medan = 67, vs. 22, ; ntensve vs. extensve), and concepton (medan = 72, ; n = 79 vs. 203, ; n = 53; ntensve vs. extensve) 6

9 500 ' ' * " 300 ~ ~ *.> ; ; -y S^ ' CO >> CD T ,,.,,, : A : I, I, " Number of cows =444, r 2 =0.02, PO.00 Wean age of calf (months) CO CO T3 C o Q. 03 O C o D CO Number of cows=232, r"=0.03, PO.00 Wean age of calf (months) Number of cows =444, r= , P= Number of cows =232, r 2 = -0.08, P=0.26 Mlk yeld per day at AI (kg) Mlk yeld per day at AI (kg) FIG. 2(A-D). Relatonshp of wean age of calves and mlk yeld of cows wth the post partum ntervals to frst Aland to concepton. were not sgnfcant (P >0.5). Cows suckled twce or less tended (P = 0.0) to have shorter ntervals between calvng and frst servce (medan = 68, ; n = 62), and concepton (medan = 67, ; n = 95) than those suckled several tmes daly; medan postpartum nterval to frst servce was 203 (3-44; n = 282) and to concepton was 202 (40-44; n = 37). The concepton rate of once or twce suckled cows (53%) tended to be hgher (P = 0.09) than those suckled several tmes daly (42%). Cows served between 5 and 8 h after beng detected n oestrus conceved at a hgher rate (49%; n = 366) than those served between 9 and 32 h (32%; n = 78; P <0.0). Cows detected n oestrus on the bass of mountng actvty, bellowng and gental dscharge or restlessness conceved at a rate of 49% (n = 245), 45% (n = 6) and 32% (n = 38), respectvely; the dfferences between groups were not sgnfcant (P =). Concepton rate was hgher (P <0.05) n cows wth marked 7

10 vulvar swellng (53%; n = 24) than those wth slght or mperceptble swellng (38%; n = 203). Cows wth clear, turbd and purulent or no gental dscharge conceved at a rate of 5% (n = 335), 43% (n = 35) and 27% (n = 74), respectvely; the dfferences between gental dscharge-groups were sgnfcant (P <0.05). Cows wth marked uterne tone at AI conceved at a hgher (P <0.00) rate (54%; n = 284) than those wth slght or mperceptble uterne tone (33%; n = 60). Dependng on AI techncans, the concepton rate vared from 3.3% to 54%; however, the dfferences between techncans were not sgnfcant (P = 0.06). Cows calved durng uly to September took a shorter tme to receve frst postpartum AI than those calved n March (Fgure 3A; P <0.05). The month of calvng as man effect on the nterval between calvng and concepton was sgnfcant (P <0.05); however, after Bonferron Adustment, the dfferences n post partum nterval to concepton between months were not sgnfcant (Fgure 3B; P >0.0). Smlarly, the month of AI as man effect appeared sgnfcant on the ntervals from calvng to frst servce (P <0.0) and to concepton (P <0.05) but after Bonferron adustment, the dfferences between months were not sgnfcant (Fgure 3C, D; P >0.05) CD a 0 2 Month of Calvng (2 = February, 2 = December) 03 2 O "S. CD o co o T3 CO CO c s Month of Calvng (2 = February, 2 = December) CD CD CD O "TO 00 - CD Month of AI (2 = February, 2 = December) Month of AI (2 = February, 2 = December) FIG. 3. (A-D) Effect of month of calvng and month ofaion the post partum ntervals to frst AI and to concepton 8

11 3.3. Investgaton 3. Evaluaton of semen and classfcaton of bulls Breed of bulls nfluenced the frst servce concepton rate (P O.00). Fresan bulls had hgher (P O.0) fertlty (56.5%, n = 84), than dd Fresan x local (43.9%, n = 4) and Sahwal x others (34.9%, n = 46). The fertlty of good bulls (52.6%, n = 266) was hgher (P O.00) than that of poor bulls (35.8%, n = 65). Thrteen nsemnatons were made wth semen of bulls wth ncomplete nformaton and therefore remaned unclassfed. The use of frozen semen resulted n hgher (P O.00) concepton rate (54.9%, n = 224) than dd the chlled semen (37.3%, n = 220). The mported frozen semen yelded hgher (P <0.00) concepton rate (57.5%, n = 74) than dd the locally produced semen (38.9%, n = 270), rrespectve of frozen or chlled-preserved. The days of chllng and sperm motlty of preserved semen dd not nfluence the concepton rate (P >0.50). The use of and 2 days chlled semen resulted n 37.2% (n = 29) and 37.9% (n = 9) concepton, respectvely. The sperm motlty of <50%, 50-60% and >60-70% resulted n concepton rates of 49.2% (n = 63), 50.0% (n = 86) and 44.4% (n = 295), respectvely (ns). The total spermatozoa per cow dose nfluenced the concepton rate (P <0.00). A total of 5 mllon or less spermatozoa resulted n lower (P <0.05) concepton rate (27.0%, n = 00) than dd 6-20 mllon (44.0%, n = 34), 2-25 mllon (58.3%, n = 08) and mllon spermatozoa (54.9%, n = 02) Investgaton 4: Accuracy of oestrus detecton studed by usng radommunoassay of progesterone n mlk The cows studed were 3-5 (medan = 6.0) years old and to 8 (medan = 3.0) partes. At calvng they weghed (medan = 270) kg and scored body condton.5 to 3.5 (medan = 2.5). The ntervals after calvng to the frst detected oestrus and to the ntaton of luteal actvty were (medan = 5; n = 82) and (medan = 08, n = 64), respectvely. The estmates of progesterone concentraton at frst rse vared from 0.5 to 9.3 (medan = 2.4). Farmers mssed detectng an oestrus (false negatve) on to 3 (medan =.0) occasons postpartum. The proporton of cows accurately detected n oestrus was 30%. Another 30% were stated to be n oestrus when they were not (false postve) and 40% were not detected when they were n oestrus. The ntervals to the ntaton of frst postpartum luteal actvty were examned n these cows. Those wth BCS 3.5 or more at calvng needed fewer days to ntate luteal actvty than ther counterparts havng BCS 3.0 or less (Fgure 4A, B; P <0.05). Cows suckled once or twce daly requred fewer days (medan = 95, ; n = 28) than the cows suckled several tmes (medan = 27, ; n = 36) (P <0.05). The medan nterval to frst postpartum oestrus was prolonged by 33.5 days due to farmers' nablty to detect the oestrous cows (P <0.05). Cows mssng detecton of oestrus had lower mlk progesterone concentraton at frst luteal actvty (medan =.8, ; n = 34) than those detected accurately n oestrus (medan = 4.2, ; n = 26) (P <0.05). A postve relatonshp appeared between the estmated progesterone concentraton durng the frst peak and the duraton of postpartum acyclcty (r = 0.284, n = 64, P <0.05). The age of the cows and ther partes dd not nfluence the ntaton of luteal actvty (P >0.50). The effects of body weght at calvng, purpose of rearng cows (dary vs. dary + draught) and breed of cows on the ntaton of luteal actvty were not sgnfcant 3.5. Investgaton 5. Evaluaton of frozen semen from producton to nsemnaton The percentage of motle spermatozoa vared from 37.9 ± 5.8 to 65.4 ± 6.6, dependng on the occasons when evaluaton was made. The results are shown n Fgure 5. The sperm motlty dropped due to dluton (P <0.05), chllng (P <0.05), freezng (P <0.00) and storng 3 months n the Bull Staton (P <0.0). Three months storage n the Dstrct AI Centre also reduced (P <0.05) sperm motlty. Transportaton of semen from the Dstrct AI Centre to the Sub Centre caused further reducton (P <0.05) n the sperm motlty. 9

12 CO 5* 400 B - 0 CO Q. To CD D) CD o 300 h I * * BCS of the cow at calvng (-5 scale) I A : BCS of the cow at frst luteal phase (-5 scale) FIG. 4. (A, B) Effect of body condton at calvng and at frst luteal phase on the post partum nterval to the ntaton of luteal actvty. 80 I I I o ' ' ' ' ' ' ' Occasons of semen evaluaton FIG. 5. Sperm motlty n fresh eaculate, and at dfferent stages of semen freezng, storage, transportaton and preparaton for AI. = fresh eaculate (65 ± 7%), 2 = dluted wth trs-egg yolk medum (62 ± 6%), 3 = dluted wth trs-egg yolk-glycerol medum, cooled down to 4 C and equlbrated for 4 h (58 ± 8%), 4 = frozen-thawed semen (48 ± 8%), 5 = thawed after week of freezng (47 ± 7%), 6 = transported from the Bull Staton to the Dstrct AI Centre (44 ± 8%), 7 = prepared for AI n the Dstrct AI Centre (4 ± 6%), 8 = stored 3 months at the Bull Staton (42 ± 6%), 9 = stored 3 months n the Dstrct AI Centre (40 ± 7%), 0 = transportedfrom the Dstrct AI Centre to a Sub Centre (38 ± 8%) and = prepared for AI n the Sub Centre (38 ± 6%). Note the values n the parenthess are the mean ± SD of 5 observatons. 20

13 The percentage of normal spermatozoa wth respect to the acrosome, mdpece and tal were between 84.6 ± 4.8 and 90.0 ± 5.2 dependng on the occasons of samplng; the percentage dropped (P <0.0) due to freezng semen (Fgure 6). 00 O Q N u % % 90h en o P 80 4 " \-- X. y~ $ 4 ~ PH O 70, l Occasons of semen evaluaton FIG. 6. The proporton of spermatozoa wth normal acrosome, mdpece and tal n fresh eaculates, and at dfferent stages of freezng, storage, transportaton and preparaton for AI. = fresh eaculate (90 ± 5%), 2 = dluted wth trs-egg yolk medum (89 ± 7%), 3 = dluted wth trs-egg yolk-glycerol medum, cooled down to 4 C and equlbrated for 4 h (84 ± 6%), 4 = frozen-thawed semen (84 ± 7%), 5 = thawed after week of freezng (85 ± 5%), 6 = transported from the Bull Staton to the Dstrct AI Centre (85 ± 5%), 7 = prepared for AI n the Dstrct AI Centre (86 ± 5%), 8 = stored 3 months at the Bull Staton (85 ± 5%), 9 = stored 3 months n the Dstrct AI Centre (85 ± 5%), 0 = transported from the Dstrct AI Centre to a Sub Centre (85 ± 4%) and = prepared for AI n the Sub Centre (85 ± 5%). Note the values n the parenthess are the mean ± SD of 5 observatons. 4. DISCUSSION 4.. Investgaton. Use of radommunoassay of progesterone n mlk to survey some reproductve characterstcs of cows n an AI programme Mlk progesterone data based on three samples (Day 0, Day 9-3 and Day 2-24) helped make a clear decson n 82.5% cows about the pregnancy. If only the pregnancy dagnoss s concerned, mlk progesterone concentraton on the day of AI and on Day 2-24 wll gve the same results wthout compromsng the accuracy. Mlk progesterone concentraton on Day 0 and Day 9-3 together not only dentfed the naccurate oestrus detecton but also gave a clear dea about the cyclcal status of 2

14 the anmals. Progesterone concentraton on Day 0 can only ndcate whether or not AI was done n a cow wth functonal luteal tssues n the ovary. In the case where veternary servces are not avalable or are too expensve, mlk progesterone concentratons on the day of AI and on Day 2-24 can be used to nterpret pregnancy results wth more than 80% accuracy; dagnoss of non-pregnancy was hghly accurate Investgaton 2. Reproductve effcency of cows under AI programmes The man fndngs were the prolonged ntervals from calvng to frst servce and to concepton. Poor nutrton condton durng calvng and thereafter, frequent sucklng of long duraton because of no practce of weanng, use of cows n draught power, hgh proporton of naccurate oestrus detecton are, among others, mportant determnants of the calvng to frst servce nterval. The local cows seem to have an nherent tendency towards long nterval from calvng to frst servce; ths may not always get corrected by cross breedng wth Fresan cattle. The system and purpose of rearng cows, BCS at AI, mlk producton at AI, nterval between oestrus and AI, degrees of vulvar swellng, nature of gental dscharge and the toncty of uterus at AI sgnfcantly nfluenced the frst servce concepton rates. Long nterval to concepton after calvng not only results n fewer calves durng the lfetme of a cow and hence hnders antcpated genetc gan but also reduces the use of AI n the ndustry. When the number of AI's per techncan ncreases, costs per cows are reduced. cost s one of the mportant prerequstes for farmers' acceptance of AI [5]. The favourable effects of good body condton of cows on the nterval to concepton demonstrated here have also been shown by other workers [6, 7, 8, 9]. Good body condton of cows s only mantaned f the negatve energy balance due to producton of mlk s covered by adequate nutrton and management [6, 20, 2]. The longer ntervals to frst servce and concepton n dary + draught cows than ther dary counterparts, evdent from our results, further supports the sgnfcant role of post partum energy balance on the reproductve events. However, the effect of chronc stress on the cow caused by draught work should not be overlooked [22]. Stress prevents cows from showng characterstcs behavoural and clncal sgns of oestrus [23]. Well developed oestrous sgns, lke marked vulvar swellng, clear, strngy mucus dscharge and marked toncty of the uterus proved n ths study to have favourable effects on concepton rate. The results of ths study demonstrated that the cows calved durng uly to September took shorter tme to receve the frst postpartum AI and to conceve than those calved durng December to March, although the dfferences were not always sgnfcant. The study area experences hot and humd clmate durng Aprl to September; green forages are mostly avalable durng uly to September. In contrast, the clmatc condtons durng December to March n Bangladesh are cold and dry. Durng December-March, the cultvable lands are occuped by crop under rrgaton programme and the barren lands are too dry to support growth of grass for communal grazng. The nteractons between clmatc factors, nutrtonal factors and genetc factors n causng reproductve neffcency are complex. Further work s needed on the specfc effects of green forage and on alternatve methods of nutrtonal management. In contrast to the data on recognzed dary breeds [24], our results ndcated a postve relatonshp between mlk yeld at AI and the ntervals to frst postpartum servce and concepton, although ths was not statstcally sgnfcant. The stuaton can be explaned by the fact that dary + draught cows yeldng less mlk proved to have longer postpartum ntervals to the frst servce and concepton. The mlk producton n ths study has been recorded on the day of nsemnaton. It s lkely that cows wth prolonged postpartum ntervals due to other reasons were about to dry off and consequently beng consdered as low-yeldng. Also t s usual that hgh yeldng cows are pad more attenton, fed well and reared under good management; all these have favourable effects on the ntervals to the frst servce and concepton. The latter stuaton s supported by the fact that hgh yeldng cows n our studes conceved at a hgher rate than ther low producng counterparts. It appears n that the overall ntervals (medan) from calvng-to-frst servce and to concepton are smlar. Ths s because most of the cows receved AI only on frst post partum oestrus and then dd not receve further nsemnatons even though they dd not conceve on the frst servce. 22

15 4.3. Investgaton 3. Evaluaton of semen and gradaton of bulls The present study ndcated that good and poor bulls as classfed on the bass of semen evaluaton data dffered wth respect to the concepton rate n artfcal nsemnaton. The crtera to udge a bull as good or poor were set accordng to the motlty, total number and morphologcal normalty of the spermatozoa. The motlty tself s a flagellar functon of spermatozoa and n accordance wth Soderqust [25], the present nvestgatons dd not fnd any relatonshp between sperm motlty and fertlty. Motle spermatozoa can be morphologcally abnormal [26]. The proporton of morphologcally abnormal spermatozoa n semen correlates negatvely wth fertlty results [25, 26, 27, 28, 29]. Spermatozoal number per cow dose sgnfcantly nfluenced the concepton rate n ths study. Ths s n accord wth the mportance of the number of functonally normal spermatozoa as a determnant of fertlty [30]. It also takes nto account Soderqust's recommendaton [25] that semen parameters need to be defned quanttatvely rather than qualtatvely to be able to predct the fertlty. Usng a combnaton of semen characters, as was done here, has also been recommended n attempts to evaluate semen to predct fertlty [3, 32]. Innate fertlty of the bull combned wth optmal sperm numbers are also mportant n determnng fertlty n an AI programme [30]. The sgnfcant effect of bull-breed on concepton rate found n the present study s further confrmaton of ths. The classfcaton used here dscrmnated to some extend between bulls lkely to have hgher or lower fertlty. Further work s warranted n rankng bulls n more categores under condtons where greater numbers of cows wth confrmed fertlty results can be used. The data of the present study reveal that cows nsemnated wth frozen semen conceved at a hgher rate than those nsemnated wth chlled semen. Ths s n contrast wth some earler reports where the concepton rate after nsemnaton wth chlled semen was hgher than that obtaned after nsemnaton wth frozen semen, provded the preservaton temperature remaned constant [33, 34, 35, 36]. The low concepton rate after nsemnaton wth chlled semen n ths study could be due to dffcultes n mantanng constant temperature durng transportaton and storage of chlled semen Investgaton 4: Accuracy of oestrus detecton studed by usng radommunoassay of progesterone n mlk Falure to detect oestrus (false negatve) and false determnaton of oestrus (false postve) are common problems n AI of cows n ntensve farmng [5, 37, 38, 39]. In accordance wth our results, false negatve and false postve categores of oestrus detecton have been found by others to be as hgh as 30 to 50% [5, 37, 39] and 7 to 30% [38], respectvely. The false postve oestrus detecton was lower n Investgaton (2%) than that n Investgaton 3 (30%). In Investgaton, the cows were recorded at the AI centres. Frstly, ths means, farmers made a rgd decson about the oestrus of the cow. Secondly, the regstraton of cows by the AI techncans n Investgaton rases the queston as to whether they made any selecton n favour of good oestrus sgn to prove ther good performance. Thrdly, n Investgaton 4, to avod our frequent vsts, some farmers mght have made ntentonal false report about the oestrus. Because cows n the false negatve category are not nsemnated and cows n the false postve category are unlkely to get pregnant n that cycle, the calvng to concepton nterval s ncreased n all cases. Cows wth good body condton at calvng and thereafter ntates postpartum cyclcty earler than those n poor body condton. Ths ndcates the mportance of good nutrtonal management of pregnant cows to cover the negatve energy balance due to growth of foetus. As shown n Investgaton 2, early ntaton of postpartum cyclcty wll reduce the nterval from calvng to frst servce and to concepton. The adverse effect of the duraton and frequency of sucklng on the ntaton of postpartum cyclcty as ndcated by our results are n accordance wth other reports [40]. Also n Investgaton 2, frequent sucklng tended to ncrease the ntervals from calvng to frst servce and to concepton. Controlled sucklng for a restrcted perod favours postpartum reproducton n cows [4]. The results of the present nvestgatons do not clarfy whether prolonged sucklng lengthens the onset of postpartum ovaran actvty or the sucklng contnues because the cows do not dry off snce they are 23

16 not pregnant. However, n commercal dary farmng, controlled weanng should be practced to dentfy the cows wth an nherent tendency to reman acyclc rrespectve of sucklng and nutrton management. The low concentraton of progesterone at the frst postpartum rse of false negatve cows rases queston as to whether ther oestrous cycle was regular or not. There are reports of short oestrous cycle [7, 42] and transent rses n progesterone before the onset of usual ovaran actvty [7, 43,44, 45]. However, one should nterpret such data cautously, especally when progesterone profles are montored n less frequently sampled mlk as n the case of our nvestgatons. None the less nadequate lutenzaton often results from defectve follcular development due to lack of LHreceptor actvty [43]. There are also clams about an organzng effect of transent progesterone rse on the ovaran-ptutary-hypothalamc axs to facltate the re-establshment of regular oestrus cycle [43, 44] Investgaton 5. Evaluaton of frozen semen from producton to nsemnaton The sperm motlty dropped sgnfcantly due to dluton of fresh semen, chllng, freezng, storng n the bull staton and n the Dstrct AI Centre and transportaton from the Dstrct AI Centre to Sub Centre. Smlarly, the proporton of normal spermatozoa sgnfcantly reduced due to freezng of semen. As a result, the cow receved semen wth 38% and 40% average sperm motlty n the Dstrct AI Centre and Sub Centre, respectvely. Ths means, the cow receves 2.0 mllon or less motle spermatozoa, gven 30 mllon total spermatozoa per cow dose. The mportance of the number of spermatozoa per cow dose was evdent n Investgaton 3. The freezng protocol, and handlng of frozen semen durng transportaton and management durng storage need to be mproved to ensure at least 50% sperm motlty at the tme of AI, gven mllon total spermatozoa per cow dose. 5. CONCLUSIONS It appears that determnaton of progesterone concentraton n mlk on the day of AI and on Day 2-24 s a good means to make decson on pregnancy results by dagnosng the non-pregnant state wth hgh accuracy. Prolonged postpartum ntervals to the ntaton of ovaran actvty, frst servce and concepton, and low concepton rate are the maor constrants lmtng the development of cattle by AI n Bangladesh. The nutrton condton of the cow, duraton and frequency of sucklng, use of cows n draught power and accuracy of heat detecton are, among others, the mportant determnants of the ntervals to the ntaton of the ovaran actvty, and to the frst servce and concepton. The classfcaton of bulls nto good and poor based on semen evaluaton data dscrmnated to some extend between bulls lkely to have hgher or lower fertlty. The end-users n Bangladesh receve frozen semen wth 2 mllon or less motle spermatozoa per cow dose; ths needs to be mproved to acheve good fertlty. From these studes several areas emerge as mportant for future work. The progesterone assay and data analyss system used here should be ntroduced as a servce to farmers to assst n reproductve management of ther farms. Improvng nutrton wll clearly beneft the cows' reproductve effcency. Economc studes should be carred out to formulate cost effectve strateges, especally for small farmers. The montorng of bulls and semen should be contnued. There s a need to mprove the producton and handlng of semen to acheve hgh qualty at the pont of nsemnaton. Farmers should have better tranng on oestrus detecton. Further tranng and workshops should be nsttuted for AI techncans to acheve a more unform and hgher fertlty results. ACKNOWLEDGEMENTS The proect was funded by the ont FAO/IAEA Dvson, Venna, Austra (Research Contract No. 8566/RB). We thank the Department of Lvestock Servces of Bangladesh for gvng us access to ts AI actvtes for the completon of ths study. 24

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