Gastrointestinal nematode (GIN) infections cause severe production losses in small ruminants, particularly in sheep, worldwide. Many sheep breeds showed resistance to GIN infection and thus breeding of sheep for GIN resistance could be feasible (Bishop, 2012). Successful selection of resistant animals is an optional worm control strategy that can cope with anthelmintic resistance (Cunha et al., 2024). However, a major problem with this approach is: how can resistant sheep be selected? (Gray et al., 1992). The number, size and reproductive capacity of the parasites in the abomasum or small intestine of host, as well as faecal egg count (FEC), haemoglobin (Hb), packed cell volume (PCV), eosinophilia, levels of immunoglobulins (Ig), dag score and FAMACHA score are all indicators of resistance (Bishop, 2012; Bell et al., 2019). Selection of Resistant sheep relies largely on the use of FEC (Karrow et al., 2014). Under ideal circumstances, selection of sheep based on both FEC and PCV along with other phenotypic indicators are desirable (Vanimisetti et al., 2004). Garole sheep of West Bengal showed resistance to GIN infection based on FEC (Brahma et al., 2022). This study was conducted to identify resistant Garole sheep based on different clinico-parasitological and haemato-biochemical parameters as phenotypic indicators of GIN resistance.

Based on phenotypic traits, 60 female Garole sheep, aged 3 to 6 months, were identified from the three villages of Sundarban delta of South 24 Parganas district in West Bengal, India. Garole sheep in those villages were reared by small and marginal farmers in a unit of 2-6 animals by semi-intensive system without routine deworming.
       
Over the course of one year (March 2023 to February 2024), FEC by modified McMaster technique (Soulsby, 1982), FAMACHA score by comparing the conjunctival colour of sheep using a FAMACHA chart (Kaplan et al., 2004) and body condition score (BCS) were studied as per the procedure described by Russel (1984) and Phythian et al. (2012), with minor modifications for local adaptability for all the selected sheep. Based on FEC, FAMACHA and BCS for nine months the Garole sheep were classified as preliminary resistant (mean EPG <100) and susceptible (mean EPG >500).
       
Haemoglobin concentration, PCV% (Jain, 1993), peripheral blood eosinophil count (Dawkins et al., 1989) and total plasma protein (TP; gm/dl) using a commercial kit, were estimated and compared between resistant (n = 20) and susceptible animals (n = 20) monthly for the last 3 months of the study.
               
All the parameters for each group on different post infection days were compared by Analyse-Compare means. The significance (P value) was recorded at 5% (P<0.05) level and 1% (P<0.01) level by separate analysis of parameters (between post-infection days) using Duncan method (One-way- ANOVA). The complete statistical analyses were done with the help of Statistical Package for Social Scientist (SPSS), Windows Version 22.0.

The EPG data generally exhibits a skewed distribution and therefore it was log transformed to normalize the data. Out of 60 sheep, persistently low FEC were observed in twenty (20) Garole sheep with monthly mean log₁₀ EPG varying from 1.73 to 1.96 and those sheep were considered as preliminary resistant sheep (Fig 1). While in rest 40 Garole sheep, FEC value was not persistent during the study period and monthly mean log₁₀EPG were ranging from 2.57 to 2.88 and those animals were considered as preliminary susceptible sheep (Fig 1). The mean FEC of preliminary resistant sheep was significantly (P<0.05) lower compared to preliminary susceptible sheep during the entire study period (Fig 1).


       
Faecal egg count (FEC) is the most important and widely accepted phenotypic indicator of host resistance against GI nematodes. In the present study, twenty out of 60 sheep demonstrated resistance to GIN infection based on FEC. Many authors reported about the heritability of FEC ranging from 0.12 to 0.44 (Miller and Horohov, 2006) indicating that based on low FEC, selection of animals could be possible.
       
Haemonchus contortus is the predominant nematode parasite in small ruminants in West Bengal, India (Jas et al. 2017) and therefore, FAMACHA score was evaluated along with FEC. Mean FAMACHA score (1.73) of resistant animals was significantly (P<0.05) lower than the susceptible sheep (2.56) in the present study (Fig 2).


       
Resistant Garole sheep showed significantly (P<0.05) higher body condition score (2.55) compared to susceptible sheep (1.81) (Fig 2). Gastrointestinal nematode infection is responsible for reduced weight gain in young animal and also causes reduction in body weight in adult animals (Jas and Ghosh 2009) and thus causing lower BCS in infected animals as observed in susceptible sheep of the present study. Therefore, FAMACHA score and BCS should be utilized to select resistant individuals along with FEC as non-invasive phenotypic indicators of host resistance.
       
Haemoglobin concentration (11.30 gm /dl) and PCV% (31.51%) of resistant sheep was significantly (P<0.05) higher than the susceptible Garole (Hb - 9.52 gm/dl and PCV - 26.710%) during all the three occasions (Fig 3).


       
Peripheral blood eosinophils count (91.33 per μl blood) and total plasma protein concentration (6.32 gm/dl) were significantly (P < 0.05) higher in resistant sheep compared to the peripheral eosinophil (37.49 per µl blood) and total protein (5.76 gm/dl) content of susceptible sheep (Fig 4).


       
Out of twenty preliminary resistant sheep one sheep failed to maintain all the health parameters of resistance, as  it had lower PCV, Hb and total protein. Thus, based on FEC, selection of resistant sheep was found to be 95% accurate in the present study. Meanwhile, among the 20 susceptible Garole sheep, four (4) sheep were found to maintain good production traits, such as BCS (3 or more), FAMACHA (between 1 and 2), Hb (above 11.00) and PCV (above 32%), despite having high FEC (mean EPG > 500). Therefore, those four Garole sheep may be considered resilient. Although FEC can successfully select resistant and susceptible sheep, but other haematological and production parameters can be useful to identify resilient  sheep, accounting for 20% (4 out of 20) among susceptible sheep in the present study.
       
For host resistance and / resilience against H. contortus, PCV is an important phenotypic marker and PCV has been shown as a heritable character for host resistance against H. contortus (Vanimisetti et al., 2004). In the present study, significantly (P<0.05) reduced value of Hb and PCV% was observed in susceptible sheep having comparatively higher level of infection compared to the resistant sheep.
       
Increased number of eosinophils locally or in the peripheral circulation have been reported to be correlated with host resistance against GIN infection (Shin et al., 2009) and it is also considered as an important phenotypic marker of host resistance against GIN (Ortolani et al., 2013). Significantly (P<0.05) higher peripheral eosinophils count was also observed in resistant Garole sheep during the last three months of study period. Thus, the haematological parameters such as Hb, PCV and peripheral eosinophils count could be utilized as phenotypic marker for host resistance in Garole sheep.
               
Hypoproteinaemia due to reduced plasma protein concentration is a common clinical pathology in GI nematodosis in small ruminants due to continuous draining of plasma protein through the damaged GI tract (Soulsby, 1982). Plasma protein concentration was found to be significantly (P<0.05) higher in resistant sheep compared to susceptible sheep in the present study. Hence serum protein concentration can be considered as a valuable indicator of host resilience and / resistance against GIN infection in sheep.

Results of the present study indicated that, along with FEC, clinical parameters such as FAMACHA score and BCS and haemato-biochemical parameters should also be considered as phenotypic indicators for selection of resistant sheep.

The authors declare that there is no conflict of interest.