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Diseases caused by small ruminant lentiviruses, Mycobacterium avium ssp. paratuberculosis (MAP), Schmallenberg virus, and peste des petits ruminants virus (PPR) is globally recognised as serious threats to the ruminant industry due to their potential to spread rapidly across boundaries. Despite their global distribution and negative impacts on ruminant production, there is a gap in knowledge of the current trends in their epidemiology among sheep and goat populations in Peninsular Malaysia. This study was therefore designed to fill the gap of knowledge concerning the seroprevalence and contributing factors of CAEV, paratuberculosis, SBV, and PPRV among small ruminants from selected flocks in Selangor, Negeri Sembilan, and Pahang states in Peninsular Malaysia. A cross-sectional study design was used to collect animal data and blood samples for serological assays simultaneously. The ID Screen (ID.VET, France) indirect ELISA screening tests were used to detect serum antibodies directed against CAEV/MVV (VISNAS Ver 0922), paratuberculosis (PARAS Ver 0516), SBV (SBVC Ver 1114) and PPRV (PPRC Ver 0821). There was 45.4% (95% CI = 40.74-50.74), 6.8% (95% CI = 4.66-9.69), 27.8% (95% CI = 23.35-32.77), and 2.6% (95% CI = 1.11-0.51) true seroprevalence for CAEV, paratuberculosis, SBV, and PPR, respectively. Geographical location and species were the risk factors for CAEV and paratuberculosis, while the management system and age of small ruminants were the risk factors for SBV. The present study is the first to document a large-scale seroprevalence of MAP and PPR infection among sheep and goat flocks in Peninsular Malaysia. The presence of PPRV and MAP antibodies among small ruminant flocks is signalling current or previous exposure to the pathogens or cross reactions with similar antigens. This finding further suggests the potential for future outbreaks of these devastating diseases among sheep and goats in Malaysia. The high seroprevalence of CAEV and SBV among small ruminants indicates high levels of exposure to the viruses in the environment, which is a potential threat to production., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The lipopolysaccharide (LPS) endotoxin and outer membrane protein (OMP) are among the virulence factors of Gram-negative bacteria responsible for inducing pathogenicity in the infected host. OMP and LPS occur on the outer membrane of M. haemolytica A2, the primary aetiological agent of pneumonic mannheimiosis in small ruminants. While the LPS is known to mediate Gram-negative bacterial infection by activating downstream inflammatory pathways, the potential role of OMP during inflammatory responses remained unclear. Hence, this study determined the effect of the OMP of M. haemolytica A2 on the serum concentration of pro-inflammatory cytokines and the male reproductive hormones (testosterone and Luteinizing Hormone). We randomly assigned twelve bucks to three groups (n = 4 bucks each): Group 1 was challenged with 2 mL PBS buffer (pH 7.0) intranasally; Group 2 received 2 mL of 1.2 X 10 9  CFU/mL whole M. haemolytica A2 intranasally; and Group 3 received 2 mL of OMP extract obtained from 1.2 X 10 9  CFU/mL M. haemolytica A2 intramuscularly. Serum samples collected at pre-determined intervals were used for the quantitative determination of the pro-inflammatory cytokines (IL-1β, IL-6, and TNFα) and reproductive hormones (testosterone and LH) using commercial sandwich enzyme-linked immunosorbent assay (ELISA). The serum concentration of IL1β was initially increased within the first-hour post-challenge in Groups 2 and 3, followed by a significant decrease in concentration at 21d and 35d (p < 0.05) in Group 3. Only mild fluctuations in IL-6 occurred in group 2, as opposed to the 1.7-fold rapid increase in TNFα within 2 h post-challenge before decreasing at 6 h. An increase in pro-inflammatory cytokines was accompanied by an acute febrile response of 39.5 ± 0.38 °C (p < 0.05) at 2 h and 40.1 ± 0.29 °C (p < 0.05) at 4 h in Group 2 and Group 3, respectively. Serum testosterone decreased significantly (p < 0.05) in both treatment groups but remained significantly (p > 0.05) lower than in Group 1 throughout the study. There was a moderate negative association between testosterone and IL1β (r = -0.473; p > 0.05) or TNFα (r = -0.527; p < 0.05) in Group 2. Serum LH also showed moderate negative associations with TNFα in Group 2 (r = -0.63; p < 0.05) and Group 3 (r = -0.54; p > 0.05). The results of this study demonstrated that M. haemolytica A2 and its OMP produced marked alterations in serum levels of pro-inflammatory cytokines and male reproductive hormones. The negative correlations between serum testosterone and inflammatory cytokines would suggest the potential role of OMP in causing male infertility by mediating innate inflammatory responses to suppress testosterone production in bucks., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Pneumonic mannheimiosis is a widespread respiratory bacterial disease of small ruminants caused by Mannheimia haemolytica serotype A2. The disease is known to affect the respiratory organs of infected animals, but its effect on other vital and reproductive organs has not been fully explored. Previous studies have demonstrated increased serum pro-inflammatory cytokine concentration post-challenge with M. haemolytica A2 and its LPS, indicating systemic inflammation in the host. This study determined the potential tissue changes and alterations of sperm parameters due to infection of M. haemolytica A2 and its LPS endotoxin. In this study, twelve experimental bucks were randomly assigned to three groups of four bucks each: group 1 (control group) were intranasally inoculated with 2 mL of PBS pH 7.0, group 2 received 2 mL of 1.2 × 10 9  CFU/mL M. haemolytica A2 intranasally, and group 3 received 2 mL of LPS extracted from 1.2 × 10 9  CFU/mL of M. haemolytica A2 intravenously. Semen samples were collected at pre-determined intervals using an electro-ejaculator and analysed immediately after collection. All experimental bucks were slaughtered via exsanguination on day 60 to collect their vital and reproductive organs at necropsy, and the samples were processed and analysed for histopathological changes. The current study has revealed that bucks challenged with M. haemolytica A2 and its LPS exhibited alterations in semen parameters such as motility, wave pattern, viability, and morphological abnormalities. Mild to moderate histopathological changes of the lung, liver, testis, epididymis, vas deferens, prostate, and lymph nodes were also observed in both challenged groups. Therefore, this study revealed the potential harmful effects of respiratory mannheimiosis on the reproductive organs of the infected bucks and sheds light on the expanse of systemic effects of this disease., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Previous investigations have revealed that lipopolysaccharide (LPS) endotoxin from certain Gram-negative bacteria could adversely affect the reproductive system of female animals. However, it is unknown whether LPS endotoxin of Mannheimia haemolytica serotype A2, the principal causative bacteria that cause pneumonic mannheimiosis in small ruminants, may also induce similar insidious effects. Therefore, this study aimed to investigate the effects of M. haemolytica serotype A2 and its LPS endotoxin on the responses of female gonadal hormones (progesterone and oestrogen), pro-inflammatory cytokines (interleukin-1β, interleukin-6), acute-phase proteins (haptoglobin and serum amyloid A) and cellular changes via histopathology study of female reproductive organs of the treatment does. Twelve clinically healthy, non-pregnant, crossbred does were randomly allocated into three equal groups. Group 1 was administered intranasally with 2 ml of sterile phosphate-buffered saline (PBS) and served as a negative control group. Group 2 was challenged intranasally with 2 ml of bacterial inoculum containing 10 9 colony-forming units (CFU)/ml of M. haemolytica serotype A2, while Group 3 was challenged intravenously with 2 ml of LPS endotoxin extracted from 10 9  CFU/ml of M. haemolytica serotype A2. Following that, blood samples were collected serially at pre-determined intervals for serological analyses. All does were euthanised 60 days post-challenges, and tissue samples from the ovaries, oviducts, uterine horns, uterine body, cervix and vagina were collected for histopathological study. The serological result revealed a significant increase (p < 0.05) in the mean concentrations of progesterone, oestrogen, interleukin-1β, interleukin-6, haptoglobin and serum amyloid A for both challenged groups. Histopathologically, all reproductive organs (except the cervix and vagina) from both challenged groups displayed significant cellular alterations (p < 0.05) characterised by haemorrhage and congestion, necrosis and degeneration, inflammatory cell infiltration and oedema. This study provides new information that elucidates the potential role of pneumonic mannheimiosis in the pathogenesis of female infertility amongst small ruminants., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Previous studies have shown that Mannheimia haemolytica A2 is the principal microorganism causing pneumonic mannheimiosis, a major bacterial respiratory disease among sheep and goats. The effect of this bacteria on the respiratory system is well-established. However, its effect on the reproductive physiology remains unclear. Therefore, this study aimed to determine the alterations in the level of pro-inflammatory cytokines and testosterone hormone post-inoculation with M. haemolytica serotype A2 and its lipopolysaccharide (LPS) endotoxin which were hypothesized to affect the reproductive functions of bucks. Twelve clinically healthy adult male goats were divided equally into three groups. Goats in group 1 were treated with 2 ml of sterile phosphate-buffered saline (PBS) pH 7.0 intranasally (negative control), group 2 with 2 ml of 10 9 colony-forming unit (CFU) of M. haemolytica serotype A2 intranasally (positive control), and group 3 were treated with 2 ml of lipopolysaccharide extracted from 10 9 CFU of M. haemolytica serotype A2 intravenously. Following inoculation, blood samples were collected via jugular venipuncture into plain tubes at pre-determined intervals for serum collection to determine the concentration of interleukin (IL)-1β, IL6, tumor necrosis factor (TNF)-α, and testosterone hormone by using commercial ELISA test kits. Results from this study demonstrated that the inoculation of M. haemolytica A2 and its LPS increases the concentration of pro-inflammatory cytokines but decreases the concentration of testosterone hormone in challenged animals at most time points throughout the 56 days experimental period (p < 0.05). This study suggests that the M. haemolytica A2 and its LPS could alter the concentration of pro-inflammatory cytokines and testosterone hormone, which in turn, may negatively affect the reproductive functions of bucks.

Mycoplasma ovis (formerly Eperythrozoon ovis) is an epierythrocytic parasitic bacterium of small ruminants known as haemotropic mycoplasma, which is transmitted mechanically by biting flies and contaminated instruments. Acute mycoplasmosis causes severe haemolytic anaemia and mortality in young animals. At the same time, chronic disease may produce mild anaemia and varying degrees of morbidity depending on several factors, including age, reproductive status, the plane of nutrition, immunological status and the presence of concurrent infection. Haemotropic Mycoplasma ovis is currently recognised as an emerging zoonotic pathogen which is widely distributed in the sheep and goat producing areas of tropics and subtropics, where the disease is nearly endemic. Human infection has been reported in pregnant women, immunocompromised patients and people exposed to animals and arthropods. The current diagnosis of haemoplasma relies on microscopic evaluation of Giemsa-stained blood smear and PCR. Although there are few published reports on the incidence of haemotropic Mycoplasma ovis infection of small ruminants in Malaysia, information on its prevalence, risk factors, severity and economic impacts is grossly inadequate. Therefore, a large-scale survey of small ruminant flocks is necessary to elucidate the current seroprevalence status and molecular characteristics of haemotropic M. ovis infection in Malaysia using ELISA and PCR sequencing technologies. In the future, surveillance programs, including vector forecast, quarantine, monitoring by periodic surveys and public enlightenment, will limit the internal and transboundary spread of M. ovis, enhance control efforts and mitigate production losses in Malaysia.